Biomolecular Machines

General Academics Science, Technology and Medicine
1

Welcome to the Molecular Machines

A thread to lump together all the interesting discoveries regarding the intracellular biomolecular machinery that are crucial for life to exist. Feel free to post interesting discoveries and perhaps describe the functionality of the intracellular biomolecular machines.

Intracellular biomolecular machinery include the following:

  1. DNA replication and repair machinery (replisome)
  2. DNA transcription machinery and RNA processing and translation machinery (Spliceosomes and ribosomes)
  3. Cell cycle signaling network (pRB, e2F, CDKs)
  4. Programmed cell death machinery (Apoptosis, autophagy, mitotic catastrophe)
  5. Protein processing machinery (Chaperones, ubiquitin-proteasome system)
  6. Intracellular signaling networks (protein kinases and phosphatases)
  7. Mechanical machines for intracellular shuttling of biomolecules and cellular movement (Microtubule network, kinesin, dynein)
    8 ) Energy production machines (Electron transport chain, F0F1 ATP synthase)

Sliding clamps and the clamp-loading machine.
Sliding clamps are ring-shaped proteins that some refer to as the “guardians” of the genome or others name them as the “ringmasters” of the genome.
Interestingly these clamps are structurally and functionally conserved in all branches of life and crystallographic studies have shown that they have almost superimposable three-dimensional structures, yet these components have very little sequence similarity (Figure 1) [1].

http://www.scienceforums.net/forum/picture.php?albumid=49&pictureid=362

Figure 1: Sliding clamps from the various domains of life.

The picture below is taken from the Molecular Biology Visualization of DNA video (2:14) from the freesciencelectures.com site.
Great video!

http://www.scienceforums.net/forum/picture.php?albumid=49&pictureid=369

Figure 2: Replication machinery.

The following components can be seen:
Sliding clamps (PCNA in eukaryotes): Green circular shaped
Clamp loader (RFC in eukaryotes): Blue-white component in the middle
(Figure 3: Structures of PCNA connected to RFC (front))

http://www.scienceforums.net/forum/picture.php?albumid=49&pictureid=367

Figure 4: Structures of PCNA connected to RFC (side)

(Figure 5: Structures of PCNA connected to RFC url=http://www.scienceforums.net/forum/album.php?albumid=49&pictureid=365)
Helicase: Blue (Figure 6: Helicase (front))
DNA polymerase: Dark-blue components attached to the sliding clamps
Primase: Green component attached to helicase
Leading strand: Spinning off to the right
Lagging strand: Spinning off to the top

They are not ringmasters for nothing.
Sliding clamps participate and control events that orchestrate DNA replication events in the following ways:

* Enhancement of DNA polymerase activity.
* Coordinate Okazaki fragment processing.
* Prevention of rereplication
* Translesion synthesis
* Prevents sister-chromatid recombination and also coordinates sister-chromatid cohesion
* Crucial role in mismatch repair, base excision repair, nucleotide excision repair
* Participates in chromatin assembly

Other functions include:

* Epigenetic inheritance
* Chromatin remodeling
* Controls cell cycle and cell death signaling

The true ringmasters.

Clamp loaders are another group of interesting proteins (see video and figures 3-5 above). Interestingly again, their functional and structural architecture are conserved across the three domains of life with low-level sequence similarity [2]. At the replication fork during replication, they load the sliding clamps many times onto the lagging strand (after DNA priming) and only once onto the leading strand. They also act as a bridge to connect the leading and lagging strand polymerases and the helicase. Which brings us to another interesting group of proteins; the helicases.

Helicases are also known to be ring-shaped motor proteins, typically hexamers (see figure 6) and separate double-stranded DNA into single-stranded templates for the replication machinery. Replication occurs at about 1000 base pairs per second due to the highly efficient combination of sliding clamps and the polymerases. Thus, helicases need to unwind DNA at at least that speed. Unwinding DNA too slowly and the replication machinery might break down . Unwind the DNA too fast or untimely and harmful mutations might occur as single-stranded DNA is prone to degradation and cytosine deamination.

The speed at which helicase unwinds DNA is no accident though, as it is intrinsically controlled. As helicase is bound to the lagging strand, it unwinds the leading strand in a separate direction. Applying a pulling force on the leading strand leads to a 7-fold increase in the speed of DNA unwinding by helicase [3, 4]. The highly efficient DNA polymerase/sliding clamp combination provides this controlling force on the leading strand. This forms a robust unwinding/polymerization interaction whereby polymerization controls and prevents unwanted DNA unwinding.

Altogether, the replisome machinery provide a robust way for DNA replication to prevent unnecessary DNA damage and mutation.

References.

  1. Vivona JB, Kelman Z. The diverse spectrum of sliding clamp interacting proteins. FEBS Lett. 2003 Jul 10;546(2-3):167-72.
  2. Jeruzalmi D, O’Donnell M, Kuriyan J. Clamp loaders and sliding clamps. Curr Opin Struct Biol. 2002 Apr;12(2):217-24.
  3. Ha T. Need for speed: mechanical regulation of a replicative helicase. Cell. 2007 Jun 29;129(7):1249-50.
  4. Johnson DS, Bai L, Smith BY, Patel SS, Wang MD. Single-molecule studies reveal dynamics of DNA unwinding by the ring-shaped T7 helicase. Cell. 2007 Jun 29;129(7):1299-309.
2

The bc1-complex for electron transfer from dihydroubiquinone to cytochrome c through the Q-cycle.

The bc1-like complexes (Complex III in mitochondria) play a central role in the electron transport chains of respiratory and photosynthetic machinery.

Their function is to carry out a sequence of electron and proton transfer reactions to generate a trans-membrane proton motive force that supplies the energy for ATP synthesizing utilizing the ATP synthase (excellent video, funny clip:p) machinery. Protons and electrons are supplied by dihydroubiquinone which in turn is generated by complexes I and II of the electron transport chain.

How do the bc1-like complexes carry out their function?
First the structure:

http://mybroadband.co.za/vb/picture.php?albumid=74&pictureid=347

The cyt bc1-complex contains two separate redox chains; High potential and low potential.
The high-potential chain connects the Qo-binding site with the cyt c1 through the Rieske Iron-sulphur-protein (RISP). The RISP is situated on a rotateable arm that is able to connect the cyt c1 component with the Qo-binging site.
The low potential chain connects the Qo-site with the Q1-site through the cyt BL and Cyt BH complexes.

Now the mechanism. A bifurcated electron transfer mechanism:

http://mybroadband.co.za/vb/picture.php?albumid=74&pictureid=401

  1. The lipid-soluble dihydroubiquinone molecule binds at the Qo-site and liberates one proton into the intermembrane space and in the process forms a semiubiquinone radical.
  2. The RISP swings around to receives an electron from the semiubiquinone and donates it to cyt c1 which in turn donates it to cytochrome c. Cytochrome c plays its part in energy transfer to complex IV of the electron transfer chain.
  3. A second proton is liberates into the intermembrane space and an electron is donated to the low potential chain, resulting in the formation of ubiquinone
  4. At the Q1-site the electron is donated to ubiquinone to form semiubiquinone, while a proton is donated from the mitochondrial matrix.
  5. In order for the formation of dihydroubiquinone at the Q1-site, two dihydroubiquinones must bins at the Qo-site.
  6. Thus the end result is the formation of 1 dihydroubiquinone, 2 quinones, 4 intermembrane protons and 2 ferrocyrochrome c proteins and loss of 2 mitochondrial matrix molecules after the binding of 2 dihydroubiquinones at the Qo-site.

That is the basic general mechanism, however research is ongoing into how bypass reactions are avoided.
For example:
Why do the electrons flow in only one direction in the low electron transport chains?
Why aren’t both electrons donated to the high-potential chain in the first place?
Radical hypotheses have been proposed including (From Cape et al. 2006 Trends Plant Sci. 2006 Jan;11(1):46-55.):

QUOTE A complex that can either stabilize the intermediate semiubiquinone, rendering it inert and invisible through some unknown mechanism, or that can use the unprecedented tactic of destabilizing its reactive intermediates.
(ii) A kinetic ‘water-park’ that tunes reaction activation enthalpies or entropies to route ‘water’ (electron) flow into productive channels.
(iii) A nano-machine that gates the electron and proton transfer reactions of semiubiquinone according to its recognition of the different redox and/or conformational states of the complex.
(iv) An extraordinary, and unprecedented, double concerted oxidation of dihydroubiquinone that simultaneously distributes two electrons and at least one proton between at least three different acceptors.
[/quote]
Options II and III do not exclude the possibility of quantum mechanics and coulombic interactions playing a role.

All-in-all a brilliant solution for a bifurcated electron transfer mechanism in order to generate a proton motive force from dihydroubiquinone.

Interestingly, the intermediate (semiubiquinone) generated at the Qo-site is believed to be a major contributor to the formation of reactive oxygen species by donating it’s free electron to oxygen and thereby resulting in the formation of superoxide. Superoxide formation causes damage to various molecules including DNA, RNA, proteins and lipids.

http://mybroadband.co.za/vb/picture.php?albumid=74&pictureid=399

Semiubiquinone

Paradoxically though, reactive oxygen specie generation at the Qo-site as a result of semiubiquinone formation is increased during periods of hypoxia (low oxygen). Hypoxia is a major initiator of cancerous growth because it activates various pro-growth signaling pathways. Hypoxia in cells usually occur as a result of poor circulation and delivery of oxygen. Obesity, lack of exercise and poor diet all contribute to these circumstances.

Thus, the bc1-complexes connects bad health choices with higher incidences of cancers and other mitochondrially related diseases through reactive oxygen species formation as a result of hypoxic conditions within various systems of the body.

Exercising and eating right are thus good for oiling your biomolecular machines…

3

Do your threads breed?

https://richarddawkins.net/forum/viewtopic.php?f=4&t=60319

ID evangelism?

4

A little more information about nuclear pores:
Video and a few images:

http://www2.uic.edu/~klierl2/images/full/nuclear_pore.png

http://www.rockefeller.edu/labheads/chait/images/nuclear_pore_complex.jpg

http://learn.genetics.utah.edu/content/begin/cells/membranes/images/NuclearPore.jpg

And a few recent discoveries:
First Detailed Map Of Nuclear Pore Complex Made
New Model Of A Nuclear Pore Complex Is Based On Crystal Structure Of Its Key Component
3-D Structure Of Key Nuclear Pore Building Block Identified

[QUOTE]In new research, scientists have for the first time glimpsed in three dimensions an entire subcomplex of the NPC; it’s the key building block of this little understood and evolutionarily ancient structure, an innovation fundamental to the development of nearly all multicellular life on earth.

The findings, by Martin Kampmann, a graduate student in John D. Rockefeller Jr. Professor Günter Blobel’s Laboratory of Cell Biology, add details to an unfolding picture of cellular evolution that shows a common architecture for the NPC and the vehicles that transport material between different parts of the cell, called coated vesicles. As early as 1980, Blobel proposed that internal membranes of cells – such as those encompassing the nucleus and vesicles – evolved from folds or invaginations of the outer cell membrane.

Rockefeller scientists Brian Chait and Michael Rout suggested in a 2004 paper in PLoS Biology that both the NPC and vesicle coats, which contain similar protein folds, evolved from ancient membrane-coating proteins that stabilized these primordial internal membranes. “So far, it’s been unclear how these ancient folds work in the nuclear pore complex”, Kampmann says. “Now we can see that the α-solenoid folds form long, flexible arms and hinges that end in the more compact, globular β-propellers. The same architectural principle is found in clathrin, a common component of vesicle coats.”

In research to be published online June 7 in Nature Structural & Molecular Biology, Kampmann isolated and purified samples of the most fundamental building block of the NPC known as the Nup84 complex, which is composed of seven proteins. The entire NPC – enormous by molecular standards – consists of 30 different kinds of proteins. Focusing on the Nup84 complex, Kampmann used an electron microscope (EM) to take thousands of images of the complex in different states or conformations, which could reflect a role in the expansion and contraction thought to facilitate the passage of various sized molecules through the NPC. By computationally averaging these many different views, he reconstructed the first three-dimensional models of the Nup84 complex. Finally, based on prior work in the Blobel lab using X-ray crystallography to determine the exact atomic structure of individual proteins in the Nup84 complex, he plugged these proteins snugly into the EM structure.

“Because the nuclear pore complex is probably too big and flexible to determine its entire atomic structure by X-ray crystallography, I think this three-dimensional EM approach could be a big help in solving the whole thing,” Kampmann says. “It allows us to put the crystal structures that we do have in context.” Kampmann is applying the EM approach to other subunits in hopes of fleshing out the overall picture of one of the most mysterious machines in molecular biology. “Martin’s data represent an important advance toward piecing together the structure of the NPC,” Blobel says.

Given the central role of the nuclear pore complex in the most basic cell processes, defects in its assembly, structure and function can have lethal consequences. Its proteins have been associated with viral infection, primary biliary cirrhosis and cancer. An understanding of how the complex works could lead to treatments for these diseases, and also reveal the evolutionary coup that led to the gene-protecting structure found in every cell more complicated than the simplest single-celled microorganisms: the nucleus.
[/quote]

5

More efficient biomolecular machines!

DNA ‘Sloppier Copier’ Surprisingly Efficient: Three Major Puzzles About Famous Enzyme Solved

ScienceDaily (July 15, 2009) — The "sloppier copier" discovered by USC biologists is also the best sixth man in the DNA repair game, an article in the journal Nature shows.
T[B]he enzyme known as DNA polymerase V (pol V) comes in when a cell's DNA is reeling from radiation damage or other serious blows. Pol V copies the damaged DNA as best it can – saving the life of the bacterial cell at the cost of adding hundreds of random mutations. [/B] [U]The July 16 Nature study reveals pol V's key attributes: economy of motion and quickness to engage[/U].

The study also solves two other stubborn mysteries about the mechanics of DNA repair: the exact composition of the active form of pol V and the crucial role of a protein filament, known as RecA*, that is always present around DNA repair sites, but was never shown to be directly involved.

The three findings together describe an exquisitely efficient process.

“It’s a beautiful mechanism for how cells conserve energy,” said first author Qingfei Jiang, a graduate student of senior author Myron Goodman, professor of biological sciences and chemistry at USC College.

Cells multiply by division, which starts with the copying of DNA. Pol V kicks in when a section of damaged DNA baffles the enzymes normally involved in copying.

In experiments with E. coli, Jiang and Goodman showed that the activation signal for pol V is the transfer to the enzyme of two key molecules from RecA*.

RecA* is a nucleoprotein filament: a long line of proteins bound to single-stranded DNA. The molecules that RecA* transfers to pol V are ATP, the energy factory of the cell, and a single RecA* protein among the many that make up the filament.

The copying of damaged DNA is formally called “translesion synthesis,” or TLS.

“What is RecA* doing?” had been a vexing question in the field for two decades, since the discovery that the filament was necessary for DNA repair. No one, however, could figure out why.

Goodman’s group showed that the role of RecA* is limited but direct: It is needed to donate molecules to activate pol V, but it does not participate in damage-induced DNA copying and does not even need to be next to the repair site.

Instead, RecA* acts as a fuel station to put pol V to action.

With the two extra molecules attached, pol V copies the damaged DNA. As soon as it reaches the end of the damaged section, it falls off and immediately deactivates.

Pol V then waits to be called again.

In addition to saving energy, the process prevents the mistake-prone copier from trying to “repair” normal DNA.

“All the other DNA polymerases [enzymes], when they copy DNA, they go first from one and then to another DNA and copy it. Not this baby. It has to be reactivated,” Goodman said.

“It’s a utility player. It’s the guy who does the tough jobs.”

He added that the discovery “explains one of the key ways that you get mutations when you damage DNA.”

Human cells use similar enzymes, Goodman said.

The study of mutations holds fundamental relevance for medicine, evolutionary biology, aging research and other fields.

Goodman’s research group discovered pol V in 1999. The “sloppier copier” nickname, coined by USC science writer Eric Mankin, has since been adopted widely.

At the time, Goodman described pol V as a “last-ditch cell defense” that averts death at the cost of frequent copying mistakes, which show up as mutations in the cell’s DNA.

Ironically, the sloppier copier may do more for the long-term success of the species than its accurate cousins. Some of the accidental mutations are likely to be helpful. Cells with those mutations will adapt better to their environment, and the mutations will spread through the species by natural selection.

Goodman and Jiang’s co-authors were Kiyonobu Karata and Roger Woodgate of the National Institute of Child Health and Human Development, and Michael Cox of the University of Wisconsin-Madison.

The National Institutes of Health funded the research.

6

Bacteria communicate with each… to share information of the perceived environment.

‘Rosetta Stone’ Of Bacterial Communication Discovered

ScienceDaily (July 13, 2009) — The Rosetta Stone of bacterial communication may have been found. Although they have no sensory organs, bacteria can get a good idea about what's going on in their neighborhood and communicate with each other, mainly by secreting and taking in chemicals from their surrounding environment. Even though there are millions of different kinds of bacteria with their own ways of sensing the world around them, Duke University bioengineers believe they have found a principle common to all of them.
The researchers said that a more complete understanding of communication between cells and bacteria is essential to the advancement of the new field of synthetic biology, where populations of genetically altered bacteria are "programmed" to do certain things. Such re-programmed bacterial gene circuits could see a wide variety of applications in medicine, environmental cleanup and biocomputing.

It is already known that a process known as “quorum sensing” underlies communication between bacteria. However, each type of bacteria seems to have its own quorum-sensing abilities, with tremendous variations, the researchers said.

“Quorum sensing is a cell-to-cell communication mechanism that enables bacteria to sense and respond to changes in the density of the bacteria in a given environment,” said Anand Pai, graduate student in bioengineering at Duke’s Pratt School of Engineering. “It regulates a wide variety of biological functions such as bioluminescence, virulence, nutrient foraging and cellular suicide.”

The researchers found that the total volume of bacteria in relation to the volume of their environment is a key to quorum sensing, no matter what kind of microbe is involved.

“If there are only a few cells in an area, nothing will happen,” Pai said. “If there are a lot of cells, the secreted chemicals are high in concentration, causing the cells to perform a specific action. We wanted to find out how these cells know when they have reached a quorum.”

Pai and scientist Lingchong You, assistant professor of biomedical engineering and a member of Duke’s Institute for Genome Sciences & Policy and Center for Systems Biology, have discovered what they believe is a common root among the different forms of quorum sensing. In an article in the July 2009 issue of the journal Molecular Systems Biology, they term this process “sensing potential.”

“Sensing potential is essentially the linking of an action to the number of cells and the size of their environment,” You said. "For example, a small number of cells would act differently than the same number of cells in a much larger space. No matter what type of cell or their own quorum sensing abilities, the relationship between the size of a cell and the size of its environment is the common thread we see in all quorum sensing systems.

“This analysis provides novel insights into the fundamental design of quorum sensing systems,” You said. “Also, the overall framework we defined can serve as a foundation for studying the dynamics and the evolution of quorum sensing, as well as for engineering synthetic gene circuits based on cell-to-cell communications.”

Synthetic gene circuits are carefully designed combinations of genes that can be “loaded” into bacteria or other cells to direct their actions in much the same way that a basic computer program directs a computer. Such re-programmed bacteria would exist as a synthetic ecosystem.

“Each population will synthesize a subset of enzymes that are required for the population as a whole to produce desired proteins or chemicals in a coordinated way,” You said. “We may even be able to re-engineer bacteria to deliver different types of drugs or selectively kill cancer cells”

For example, You has already gained insights into the relationship between predators and prey by creating a synthetic circuit involving two genetically altered lines of bacteria. The findings from that work helped define the effects of relative changes in populations.

The research was supported by National Institutes of Health, a David and Lucile Packard Fellowship, and a DuPont Young Professor Award.


So, these scientists are going to intelligently mimic the fundamental design of these quorum sensing systems, front-loading and re-programming them to produce medicinally relevant compound and/or enzymes. Fascinating.

7

And these molecular autonamous agents control their own movement…in groups.
By Manipulating Oxygen, Scientists Coax Bacteria Into Never-Before-Seen Solitary Wave

ScienceDaily (July 17, 2009) — Bacteria know that they are too small to make an impact individually. So they wait, they multiply, and then they engage in behaviors that are only successful when all cells participate in unison. There are hundreds of behaviors that bacteria carry out in such communities. Now researchers at Rockefeller University have discovered one that has never been observed or described before in a living system.
In research published in the May 12 issue of Physical Review Letters, Albert J. Libchaber, head of the Laboratory of Experimental Condensed Matter Physics, and his colleagues, including first author Carine Douarche, a postdoctoral associate in the lab, show that when oxygen penetrates a sample of oxygen-deprived Escherichia coli bacteria, they do something that no living community had been seen to do before: The bacteria accumulate and form a solitary propagating wave that moves with constant velocity and without changing shape. But while the front is moving, each bacterium in it isn’t moving at all.

“It’s like a soliton,” says Douarche. “A self-reinforcing solitary wave.”

Unlike the undulating pattern of an ocean wave, which flattens or topples over as it approaches the shore, a soliton is a solitary, self-sustaining wave that behaves like a particle. For example, when two solitons collide, they merge into one and then separate into two with the same shape and velocity as before the collision. The first soliton was observed in 1834 at a canal in Scotland by John Scott Russell, a scientist who was so fascinated with what he saw that he followed it on horseback for miles and then set up a 30-foot water tank in his yard where he successfully simulated it, sparking considerable controversy.

The work began when Libchaber, Douarche and their colleagues placed E. coli bacteria in a sealed square chamber and measured the oxygen concentration and the density of bacteria every two hours until the bacteria consumed all the oxygen. (Bacteria, unlike humans, don’t die when starved for oxygen, but switch to a nonmotile state from which they can be revived.) The researchers then cracked the seals of the chamber, allowing oxygen to flow in.

The result: The motionless bacteria, which had spread out uniformly, began to move; first those around the perimeter, nearest to the seals, and then those further away. A few hours later, the bacteria began to spatially segregate into two domains of moving and nonmoving bacteria and pile up into a ring at the border of low-oxygen and no-oxygen. There they formed a solitary wave that propagated slowly but steadily toward the center of the chamber without changing its shape.

The effect, which lasted for more than 15 hours and covered a considerable distance (for bacteria), could not be explained by the expression of new proteins or by the addition of energy in the system. Instead, the creation of the front depends on the dispersion of the active bacteria and on the time it takes for oxygen-starved bacteria to completely stop moving, 15 minutes. The former allows the bacteria to propagate at a constant velocity, while the latter keeps the front from changing shape.

However, a propagating front of bacteria wasn’t all that was created. “To me, the biggest surprise was that the bacteria control the flow of oxygen in the regime,” says Libchaber. “There’s a propagating front of bacteria, but there is a propagating front of oxygen, too. And the bacteria, by absorbing the oxygen, control it very precisely.”

Oxygen, Libchaber explains, is one of the fastest-diffusing molecules, moving from regions of high concentration to low concentration such that the greater the distance it needs to travel, the faster it will diffuse there. But that is not what they observed. Rather, oxygen penetrated the chamber very slowly in a linear manner. Equal time, equal distance. “This pattern is not due to biology,” says Libchaber. “It has to do with the laws of physics. And it is organized in such an elegant way that the only thing it tells us is that we have a lot to learn from bacteria.”

Through communication, even the most primitive life forms manipulate their surroundongs towards an end… survival.

8

Molecular Typesetting: How Errors Are Corrected While Proteins Are Being Built

[QUOTE]ScienceDaily (June 23, 2009) — Researchers at the Universities of Leeds and Bristol have developed a model of how errors are corrected whilst proteins are being built.
[/quote]

[QUOTE]Ensuring that proteins are built correctly is essential to the proper functioning of our bodies, but the ‘quality assurance’ mechanisms that take place during this manufacturing process are not fully understood.

“Scientists have been puzzled as to how this process makes so few mistakes”, says Dr Netta Cohen, Reader at the University of Leeds’ School of Computing.

To create a protein, the first step involves copying the relevant gene on our DNA onto a template, called RNA. This copying process is carried out by molecular machines called RNA polymerases.

“The RNA polymerase acts like an old fashioned newsprint typesetter, constructing newsprint by assembling letters one at a time. Similarly, RNA polymerase constructs RNA by reading the DNA and adding new letters to the RNA one at a time,” explains Dr Cohen.

There’s no way for the RNA polymerase to ensure that the correct letter is always incorporated at the right spot. “Statistically, we would expect to see a hundred-fold more errors than we actually do, so we know that some error correction must be happening. Otherwise, many more proteins in our bodies would malfunction,” says Dr Cohen.

Biological experiments have shown that the RNA polymerase slides both forwards and backwards along the RNA sequence it has created. What’s more, it has miniature scissors that can then cut out the last few letters of RNA.

So how are errors corrected? Intelligent typesetters would remove the last few letters when they spot an error. The new model suggests how the backward sliding stalls when passing an error, so wrong letters can be snipped off and copying can resume.

“The mechanism we’ve modelled has only recently been shown to be implicated in proofreading,” says Dr Cohen. “In fact, there is more than one identified mechanism for ensuring that genetic code is copied correctly. The challenge now is to find out – through a combination of experimental biology and modelling – which mechanism is dominant.”
[/quote]
Intelligent typesetters? Mmmm. Even bacteria have those.

9

Cells Are Like Robust Computational Systems, Scientists Report

[QUOTE]ScienceDaily (June 16, 2009) — Gene regulatory networks in cell nuclei are similar to cloud computing networks, such as Google or Yahoo!, researchers report today in the online journal Molecular Systems Biology. The similarity is that each system keeps working despite the failure of individual components, whether they are master genes or computer processors.
[/quote]

[QUOTE]This finding by an international team led by Carnegie Mellon University computational biologist Ziv Bar-Joseph helps explain not only the robustness of cells, but also some seemingly incongruent experimental results that have puzzled biologists.

"Similarities in the sequences of certain master genes allow them to back up each other to a degree we hadn’t appreciated," said Bar-Joseph, an assistant professor of computer science and machine learning and a member of Carnegie Mellon’s Ray and Stephanie Lane Center for Computational Biology.

Between 5 and 10 percent of the genes in all living species are master genes that produce proteins called transcription factors that turn all other genes on or off. Many diseases are associated with mutations in one or several of these transcription factors. However, as the new study shows, if one of these genes is lost, other “parallel” master genes with similar sequences, called paralogs, often can replace it by turning on the same set of genes.

That would explain the curious results of some experiments in organisms ranging from yeast to humans, in which researchers have recently identified the genes controlled by several master genes. Researchers have been surprised to find that when they remove one master gene at a time, almost none of the genes controlled by that master gene are de-activated.

In the current work, the Carnegie Mellon researchers and their colleagues in Israel and Spain identified the most probable backup for each master gene. They found that removing the master genes that had very similar backups had almost no noticeable effect, but when they removed master genes with less similar backups, the effect was significant. Additional experiments showed that when both the master gene and its immediate backup were removed, the effects became very noticeable, even for those genes with a similar backup gene. In one example, when the gene Pdr1 was removed, researchers found almost no decrease in activation among the genes it controls; when Pdr1 and its paralog were removed, however, 19 percent of the genes Pdr1 controls failed to activate.

“It’s extremely rare in nature that a cell would lose both a master gene and its backup, so for the most part cells are very robust machines,” said Anthony Gitter, a graduate student in Carnegie Mellon’s Computer Science Department and lead author of the Nature MSB article. “We now have reason to think of cells as robust computational devices, employing redundancy in the same way that enables large computing systems, such as Amazon, to keep operating despite the fact that servers routinely fail.”

In addition to Bar-Joseph and Gitter, the authors include Itamar Simon, Zehava Siegfried and Michael Klutstein of Hebrew University Medical School in Jerusalem, Oriol Fornes of the Municipal Institute for Medical Research in Barcelona, and Baldo Oliva of Pompeu Fabra University, also in Barcelona.

This work was supported by grants from the National Science Foundation and the National Institutes of Health. Molecular Systems Biology is a peer-reviewed journal published by Nature Publishing Group.
[/quote]
Resiliency and redundancy… signs of an optimal system.

10

Gene Transcribing Machine Takes Halting, Backsliding Trip Along The DNA

ScienceDaily (July 30, 2009) — [B]The body's nanomachines that read our genes don't run as smoothly as previously thought[/B], according to a new study by University of California, Berkeley, scientists.
Apparently things are a little more complicated...
[B]When these nanoscale protein machines encounter obstacles as they move along the DNA, they stall, often for minutes, and even backtrack as they transcribe DNA that is tightly wound to fit inside the cell's nucleus.[/B]

The findings come from delicate measurements of molecular-scale forces exerted on individual proteins that move along DNA to perform the first step of gene expression. These proteins, called RNA polymerase II (Pol II), slide along the DNA’s double helix, reading the genetic code and transcribing it into RNA, which is used as a blueprint to build proteins or as a switch to regulate other genes.

The measurements, which employed optical tweezers to grab both the polymerase and the end of a single molecule of DNA, are reported in the July 31 issue of the journal Science.

In collaboration with the laboratory of Mikhail Kashlev at the National Cancer Institute, UC Berkeley graduate students Courtney Hodges, Lacra Bintu and their advisor, UC Berkeley’s Carlos Bustamante, developed an optical tweezers assay to directly watch individual Pol II complexes as they move along single molecules of DNA. Optical tweezers use laser light to trap and follow a single polymerase in real time, revealing that it truly acts like a biological nanoscale machine as it moves along our genes.

The main obstacle to smooth operation of Pol II is the nucleosome, a bundle of eight histone proteins around which DNA wraps tightly . Tens of thousands of nucleosomes are bundled together into a chromosome, efficiently packaging six feet of DNA into a nucleus a million times smaller. The researchers were able to place a single nucleosome in front of the polymerase and then use the optical tweezers to observe what happens when the polymerase encounters this roadblock.

“For over 30 years, scientists had wondered how the polymerase responded to the nucleosome, and we were finally able to observe this process directly,” Hodges said. "People thought that the polymerase is a powerful motor that would blow through the nucleosome like a bulldozer, but it’s surprisingly delicate in its response; if anything is in the way, Pol II stops and backs up."

Bintu noted that this halting movement – 20-50 steps forward, then a couple of steps back – could be a key part of how gene expression is regulated. Nucleosomes are highly regulated by other proteins and can provide signals that control Pol II, much like a traffic light regulates street traffic, she said. Regulatory proteins may bind to the nucleosome and make the DNA unwind more easily, or could latch onto Pol II and prevent it from backsliding. Either would speed up transcription, while regulatory proteins that compact DNA and nucleosomes further slow down or even stop transcription.

Scientists have for years imagined that nucleosomes must be “loosened up” to allow for gene expression, and the authors note that their results give a more detailed, mechanistic insight into this process.

“Our study indicates that modulation of the wrapping/unwrapping equilibrium of DNA around the histone octamer constitutes the physical basis for regulation of transcription through nucleosomal DNA,” the authors wrote.

On the flip side, disturbances in nucleosome regulation could lead to disease.

“Transcription is a central point of control for gene expression, since everything from coordination of development to prevention of uncontrolled cell growth, that is, cancer, involves a highly regulated program of transcription by Pol II,” Bintu said. “When transcription goes haywire, pathologies like cancer and developmental abnormalities usually follow.”

Hodges and Bintu compare the DNA in the nucleosome to a band of sticky Velcro looped a couple of times around the histone proteins. The DNA is constantly being pushed around, however, and tends to peel off and then reattach to the histones. When the DNA is bound to the histones, Pol II cannot read it and transcription pauses. The polymerase restarts transcription only when the DNA briefly comes off the histones and, acting like a ratchet, works its way along the DNA throughout the entire nucleosome. At some point, the nucleosome leapfrogs over Pol II and the nanomachine trundles along unhindered.

The researchers also tugged on the two ends of a DNA molecule after transcription to see what had happened to the nucleosome. They found that the nucleosome was frequently ejected from the DNA because the tension prevented the DNA from forming loops that would have allowed the nucleosome to skip over Pol II.

“We found that even a very small amount of tension in the DNA – 3 to 5 piconewtons – during transcription results in Pol II removing the nucleosome from DNA like a pair of wire strippers,” Hodges said. “It’s very likely that the DNA in our bodies is very taut at some places and loose in others, so we think it’s possible that the cell uses tension in the genome to alter the dynamics of nucleosomes in certain genes.”

"These experiments give a much more dynamic picture of the nucleosome, showing that it isn’t a static bead-on-a-string but an active structure that can regulate when and how our genetic information is read," Bintu said. “This is just one single nucleosome, but it is the first step in understanding epigenetic effects that make one cell behave differently from another.”

Hodges is in the biophysics graduate group, and Bintu is a graduate student in physics. Both are part of Bustamante’s Jason L. Choy Laboratory of Single-Molecule Biophysics, named after a chemistry graduate student who died in an automobile accident in 2005. Bustamante is a professor of physics, chemistry and of molecular and cell biology, a Howard Hughes Medical Institute investigator and an affiliate of the California Institute for Quantitative Biosciences (QB3).

Welcome…

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ScienceDaily (July 17, 2009) — Bacteria know that they are too small to make an impact individually.

You’re sayin’ that bacteria can reason?

Mintaka

PS: Congrats on the full membership! ;D

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Oh, yes, he thinks cells are intelligent :stuck_out_tongue:

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Bacteria can reason and are intelligent? Can’t remember saying that so… no. Seems a bit of a stretch really, something panpsychists (aka materialists on steroids) might argue. Rather, living things demonstrate at least goal-directed behaviour towards survival.

Anyway, since you guys are such science lovers, I thought you might like this: Another code of life.

Researchers Crack ‘Splicing Code,’ Solve a Mystery Underlying Biological Complexity

ScienceDaily (May 6, 2010) — Researchers at the University of Toronto have discovered a fundamentally new view of how living cells use a limited number of genes to generate enormously complex organs such as the brain.

http://www.sciencedaily.com/images/2010/05/100505133252.jpg
Researchers at the University of Toronto have discovered a fundamentally new view of how living cells use a limited number of genes to generate enormously complex organs such as the brain. (Credit: Created by Graham Johnson of grahamj.com for HHMI Copyright 2005 and updated with Frey et al copyright 2010.)

In a paper published on May 6 in the journal Nature entitled "Deciphering the Splicing Code," a research team led by Professors Brendan Frey and Benjamin Blencowe of the University of Toronto describes how a hidden code within DNA explains one of the central mysteries of genetic research -- namely how a limited number of human genes can produce a vastly greater number of genetic messages. The discovery bridges a decade-old gap between our understanding of the genome and the activity of complex processes within cells, and could one day help predict or prevent diseases such as cancers and neurodegenerative disorders.

When the human genome was fully sequenced in 2004, approximately 20,000 genes were found. However, it was discovered that living cells use those genes to generate a much richer and more dynamic source of instructions, consisting of hundreds of thousands of genetic messages that direct most cellular activities. Frey, who has appointments in Engineering and Medicine, likens this discovery to “hearing a full orchestra playing behind a locked door, and then when you pry the door open, you discover only three or four musicians generating all that music.”

To figure out how living cells generate vast diversity in their genetic information, Frey and postdoctoral fellow Yoseph Barash developed a new computer-assisted biological analysis method that finds ‘codewords’ hidden within the genome that constitute what is referred to as a ‘splicing code’. This code contains the biological rules that are used to govern how separate parts of a genetic message copied from a gene can be spliced together in different ways to produce different genetic messages (messenger RNAs). “For example, three neurexin genes can generate over 3,000 genetic messages that help control the wiring of the brain,” says Frey.

“Previously, researchers couldn’t predict how the genetic messages would be rearranged, or spliced, within a living cell,” Frey said. “The splicing code that we discovered has been successfully used to predict how thousands of genetic messages are rearranged differently in many different tissues.” Blencowe’s group, including graduate student John Calarco, generated experimental data used to derive and test predictions from the code. “That the splicing code can make accurate predictions on such a large scale is a major step forward for the field,” says Blencowe.

Frey and Blencowe attribute the success of their project to the close collaboration between their team of talented computational and experimental biologists. “Understanding a complex biological system is like understanding a complex electronic circuit. Our team ‘reverse-engineered’ the splicing code using large-scale experimental data generated by the group,” Frey said.

Prof. Frey has appointments to the Canadian Institute for Advanced Research and the U of T’s Department of Electrical and Computer Engineering, the Banting & Best Department of Medical Research (BBDMR) and the Department of Computer Science. Prof. Blencowe works in the University’s Donnelly Centre for Cellular & Biomolecular Research and has appointments in the BBDMR and Department of Molecular Genetics

The research was supported by the Government of Canada through Genome Canada and the Ontario Genomics Institute, the Canadian Institutes of Health Research, National Cancer Institute of Canada, and Microsoft Research. Frey is an NSERC EWR Steacie Fellow and said that the fellowship was critical in freeing up resources so he could complete the project. The authors of the study are: Yoseph Barash, John A. Calarco, Weijun Gao, Qun Pan, Xinchen Wang Ofer Shai, Benjamin J. Blencowe & Brendan J. Frey.

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uhm, okay. random.

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Oh, they discovered another code:
Scientists Discover New Genetic Sub-Code

[b]ScienceDaily (Apr. 18, 2010) — Biologists and computer scientists from ETH Zurich and the Swiss Institute of Bioinformatics joined forces to chase possible sub-codes in genomic information. The study led to the identification of novel sequence biases and their role in the control of genomic expression.[/b]
A sub-code in the DNA enables scientists to know which genes are turned-on quickly and which are best expressed slowly.

Each cell of an organism contains a copy of its genome, which is a sequence of desoxyribo nucleotides, also called DNA. The cell is able to translate some of the coding sequences into different proteins, which are necessary for an organism’s growth, the repair of some tissues and the provision of energy.

For this translation work, the cell follows a decoding procedure provided by the “genetic code,” which tells what protein is made from a given sequence. The researchers from ETH and Swiss Institute of Bioinformatics (SIB) now identified a new sub-code that determines at which rate given products must be made by the cell. This information has several interesting implications. First, it provides novel insights into how the decoding machinery works. Secondly, and more pragmatically, it makes possible to read information about gene expression rates directly from genomic sequences, whereas up to now, this information could only be obtained through laborious and expensive experimental approaches, such as microarrays.

“A cell must respond very quickly to injuries such as DNA damage and to potent poisons such as arsenic. The new sub-code enables us to know which genes are turned-on quickly after these insults and which are best expressed slowly. One benefit of this study is that we now can get this information using only analysis of the coding sequence,” said Gina Cannarozzi, co-author of the study and Senior Research Associate at the Institute of Computational Science of ETH Zurich.

Insight into functioning of ribosomes

Additionally, the new sub-code provides insight into cellular processes at the molecular level. In every living cell, the translation allowing the production of proteins takes place at specialised factories, the ribosomes. The discovery of this novel sub-code will therefore also provide more information about the functioning of these ribosomes. Indeed, all the data gathered up to now indicate that these factories recycle their own components, the tRNAs, to optimize the speed of protein synthesis. This discovery of a new way to regulate translation could potentially be exploited to more efficiently produce therapeutic agents and research reagents. For example, many therapeutic agents, such as insulin, are produced by expressing a protein in a foreign host such as Escherichia coli or the yeast Saccharomyces cerevisiae. The new sub-code can be now used to rewrite the information such as to optimize in a much more rational manner the amount of product delivered by the foreign host.

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Nice post thanks Teleo. Interesting read.

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The exact mechanism of complex III of the electron transport chain is currently under investigation (as mentioned here).

Meanwhile, research into the mechanisms of electron transfer of complex I are yielding interesting results:
Nanomachines in the Powerhouse of the Cell: Architecture of the Largest Protein Complex of Cellular Respiration Elucidated

ScienceDaily (July 2, 2010) — Scientists of the University of Freiburg and the University of Frankfurt have elucidated the architecture of the largest protein complex of the cellular respiratory chain.They discovered an unknown mechanism of energy conversion in this molecular complex. The mechanism is required to utilize the energy contained in food.

http://www.sciencedaily.com/images/2010/07/100702100414.jpg

The structural model of mitochondrial complex I provides new insights in energy conversion at nanoscale. A molecular coupling device links pump modules in the membrane arm of the huge enzyme complex. (Credit: Image courtesy of Albert-Ludwigs-Universität Freiburg)

[b]After ten years of research work[/b], the x-ray crystallographic analysis of the huge and most complicated protein complex of the mitochondrial respiratory chain was successful. [b]The complex contains more than 40 different proteins[/b], marks the entry to cellular respiration and is thus also called mitochondrial complex I. The results are published in the current online-edition of the journal Science.

A detailed understanding of the function of complex I is of special medical interest. Dysfunction of the complex is implicated in several neurodegenerative diseases such as Parkinson´s disease or Alzheimer´s disease, and also with the physiological processes of biological aging, in general. The work of Prof. Carola Hunte of the Freiburg Institute for Biochemistry and Molecular Biology and the Freiburg excellence centre BIOSS (Centre for Biological Signalling Studies) in cooperation with Prof. Ulrich Brandt, Professor for Molecular Bioenergetics and member of the excellence centre „Macromolecular Complexes" and Dr. Volker Zickermann of his research group is a major step forward to this understanding.

The energy metabolism takes place in the so-called powerhouses of the cell, the mitochondria. They transduce the energy taken up as food into adenosine triphosphate, in short ATP, which is the universal energy currency of life. A chain of five complicated molecular machines in the mitochondrial membrane are responsible for the energy conversion. The production of ATP in mitochondria requires so many steps, as it is in principal a Knallgasreaction. In a laboratory experiment, hydrogen and oxygen gas would react in an explosion and the energy contained would be released as heat. In biological oxidation, the energy will be released by the membrane bound protein complexes of the respiratory chain in a controlled manner in small packages. Comparable to a fuel cell, this process generates an electrical membrane potential, which is the driving force of ATP synthesis. The total surface of all mitochondrial membranes in a human body covers about 14.000 square meter. This accounts for a daily production of about 65 kg of ATP.

The now presented structural model provides important and unexpected insights for the function of complex I. A special type of „transmission element," which is not known from any other protein, appears to be responsible for the energy transduction within the complex by mechanical nanoscale coupling. Transferred to the technical world, this could be described as a power transmission by a coupling rod, which connects for instance the wheels of a steam train. This new nano-mechanical principle will now be analysed by additional functional studies and a refined structural analysis.

Original article:
Functional Modules and Structural Basis of Conformational Coupling in Mitochondrial Complex I

Proton-pumping respiratory complex I is among the largest and most complicated membrane protein complexes. Its function is critical for efficient energy supply in aerobic cells and malfunctions are implicated in many neurodegenerative disorders. Here, we report the x-ray crystallographic analysis of mitochondrial complex I. The positions of all iron-sulfur clusters relative to the membrane arm were determined in the complete enzyme complex. The ubiquinone reduction site resides close to 30 Å above the membrane domain. The arrangement of functional modules suggests conformational coupling of redox chemistry with proton pumping and essentially excludes direct mechanisms. We suggest that a ~60 Å long helical transmission-element is critical for transducing conformational energy to proton-pumping elements in the distal module of the membrane arm.
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Is a “biomolecular machine” the same thing as an enzyme?

Mintaka

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Depends on your definition of machine and enzyme I guess.

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An enzyme is a catalyst, usually (if not always)a protein. A machine is an assembly that moves so that a mechanical job can be done. So I’m curious why the array above would be classed as a machine instead of an enzyme.

ETA: Maybe the proton pump? If protons are relocated through the membrane, I s’pose that could count as a mechanical effort. Nifty.

Mintaka