Production of biodiesel from Bacteria

Scientists edge nearer unlimited blood bank

French scientists have managed to generate red blood cells from stem cells and inject them back in to the donor. This major achievement opens up the possibility of a stem cell-based alternative to donated blood cells.

Waste Water + Bacteria = Clean Energy

For the first time, researchers have sustainably produced hydrogen gas, a potential source of clean energy, using only water and bacteria. The challenge now, scientists say, is to scale up the process to provide large amounts of hydrogen for various purposes, such as fueling vehicles or small generators.

Hydrogen may be the ultimate clean fuel because burning it—in chemical terms, reacting it with oxygen—yields only water vapor. Previously, researchers have produced hydrogen gas in microbial-powered, batterylike fuel cells, but only when they supplemented the energy produced by the bacteria with electrical energy from external sources—such as that obtained from renewable sources or burning fossil fuels, says Bruce Logan, an environmental engineer at Pennsylvania State University, University Park. Also, by using devices that contain large stretches of permeable membranes that separate salt water from fresh, scientists have tapped the voltage difference that exists between them. But those devices create only a voltage difference; they don't generate the electrical current required to produce hydrogen, Logan notes. Hydrogen atoms are formed in such devices only when electrons flow into a fluid where they can combine with hydrogen ions; those atoms in turn combine with each other to create hydrogen gas.

Bacterial gas. Using a prototype system that uses only fresh water (bottle, left), salt water (right), and a chamber where certain types of energy-generating bacteria feed on nutrients (foreground), scientists have produced hydrogen gas (collected in chamber at arrow) without using any external sources of energy.

Now, Logan and Penn State environmental engineer Younggy Kim report online this week in the Proceedings of the National Academy of Sciences that they've done something no other team has: They've successfully combined the two types of devices to generate hydrogen without any external sources of energy whatsoever. The prototype device contains two small chambers—one holding the bacteria and their nutrients, the other holding salty water where the hydrogen was produced—that are separated by five stacked cells through which the researchers circulated fresh water and salt water. Together, these stacked cells generated between 0.5 and 0.6 volts—enough, the researchers say, to enable hydrogen production in the microbial fuel cell, in which bacteria feed on acetate compounds.

For each 30 milliliters of sodium acetate solution provided for the bacteria, the device generated between 21 and 26 milliliters of hydrogen gas over the course of a day. Admittedly, this is a small volume, about four times the amount of fuel in a disposable lighter, but it's enough to prove that the hydrogen-generating concept works in the lab, the researchers contend. Although the equipment needed to produce the hydrogen is expensive, the device needs no external source of energy—and therefore no greenhouse gases are generated during the process.

The team's device "is elegantly simple, and their test results are well-explained and unambiguous," says Leonard Tender, a chemist at the U.S. Naval Research Laboratory in Washington, D.C. One of the challenges to scaling up the process, he notes, will be developing new materials for fuel cell membranes that won't quickly become clogged with the chemical byproducts of bacterial activity, which would cut down on the flow of ions that help maintain the voltage difference across the membranes. Once such hurdles are crossed, however, the process offers the intriguing possibility of using the organic matter in wastewater to generate energy, he notes.

But César Torres, a chemical engineer at Arizona State University, Tempe, suggests that the new technology isn't quite ready for full-scale production of hydrogen. "This is a simple process, but the chemistry and the components are complicated," he says. "The technology needed to design and manufacture materials needed to produce efficient, nonclogging membranes is quickly evolving, but there's still a lot of research to be done."

Another challenge to scaling up will be "keeping the bacteria happy," he notes. The key, he suggests, will be extracting much but not all the energy produced by the bacteria. Trying to use all of the energy produced by bacterial metabolism wouldn't leave enough for the microbes to grow, reproduce, and thrive.

HIV-Mode of action of NNRTIs

NNRTIs are a class of anti-HIV drugs. When one NNRTI is used in combination with other anti-HIV drugs – usually a total of 3 drugs – then this combination therapy can block the replication of HIV in a person's blood.

NNRTIs, sometimes referred to as "Non-Nucleoside Analogues" – or "non-nukes" for short – prevent healthy T-cells in the body from becoming infected with HIV.

 

When HIV infects a cell in a person's body, it copies it's own genetic code into the cell's DNA. In this way, the cell is then "programmed" to create new copies of HIV. HIV's genetic material is in the form of RNA. In order for it to infect T-cells, it must first convert its RNA into DNA. HIV's reverse transcriptase enzyme is needed to perform this process.

NNRTIs attach themselves to reverse transcriptase and prevent the enzyme from converting RNA to DNA. In turn, HIV's genetic material cannot be incorporated into the healthy genetic material of the cell, and prevents the cell from producing new virus.

DNA Helicase: Function, Structural Features And Superfamilies

DNA helicase is an enzyme that aids in DNA synthesis by 'unzipping' the two strands of a DNA helix so that DNA polymerase can access the DNA to add nucleotides and effect copying.


Many cellular processes (DNA replication, RNA transcription, DNA recombination, DNA repair, Ribosome biogenesis) involve the separation of nucleic acid strands. Helicases are often utilized to separate strands of a DNA double helix or a self-annealed RNA molecule using the energy from ATP or GTP hydrolysis. They move incrementally along one nucleic acid strand of the duplex with a directionality specific to each particular enzyme. There are many helicases (14 confirmed in E. coli, 24 in human cells) resulting from the great variety of processes in which strand separation must be catalyzed.

Function

Helicases adopt different structures and oligomerization states. Whereas DnaB-like helicases unwind DNA as donut shaped hexamers, other enzymes have been shown to be active as monomers or dimers. Recent studies showed that helicases do not merely wait passively for the fork to widen, but play an active role in forcing the fork to open, thus "it is an active unwinding motor". However, the unwinding is much faster in cells than in the test tube, so "accessory proteins are helping the helicase out by destabilizing the fork junction".

Structural features

The common function of helicases accounts for the fact that they display a certain degree of amino acid sequence homology; they all possess common sequence motifs located in the interior of their primary sequence. These are thought to be specifically involved in ATP binding, ATP hydrolysis and translocation on the nucleic acid substrate. The variable portion of the amino acid sequence is related to the specific features of each helicase.

Based on the presence of defined helicase motifs, it is possible to attribute a putative helicase activity to a given protein, though the presence of a motif does not confirm the protein as a helicase. Conserved motifs do, however, support an evolutionary homology among enzymes. Based on the presence and the form of helicase motifs, helicases have been separated in 4 superfamilies and 2 smaller families. Some members of these families are indicated, with the organism from which they are extracted, and their function.

Superfamilies

• Superfamily I: UvrD (E. coli, DNA repair), Rep (E. coli, DNA replication), PcrA (Staphylococcus aureus, Bacillus anthracis and Bacillus cereus, regulation of recombination by displacing RecA from DNA and inhibiting RecA-mediated DNA strand exchange), Dda (bacteriophage T4, replication initiation).
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• Superfamily II: RecQ (E. coli, DNA repair), eIF4A (Baker's Yeast, RNA translation), WRN (human, DNA repair), NS3 (Hepatitis C virus, replication). TRCF (Mfd) (E.coli, transcription-repair coupling factor).
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• Superfamily III: LTag (Simian Virus 40, replication), E1 (human papillomavirus, replication), Rep (Adeno-Associated Virus, replication, site-specific integration, virion packaging).
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• DnaB-like family: DnaB (E. coli, replication), gp41 (bacteriophage T4, DNA replication),T7gp4 (bacteriophage T7, DNA replication).
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• Rho-like family: Rho (E. coli, Transcription termination factor ).

Lecture on Stem Cells and the End of Aging

Human tissues vary in their ability to heal and regenerate. The nervous system has weak powers of regeneration, while the skin is quick to make new cells for repair. Mammalian muscle cells are intermediate in their ability to regenerate. Human muscle can regenerate in response to minor wounds and normal wear and tear, but humans will not grow a new bicep, for example, in response to amputation. The heart is the most important muscle in the body and yet has feeble regenerative capabilities. Research into the wholesale production of new replacement organs and limbs is in its infancy, but research into enhancing normal levels of regeneration is progressing rapidly. Recent discoveries concerning the location and characteristics of adult stem cells and the signals that wounded tissue produces to activate stem cells have increased our understanding of regeneration. Insulin-like growth factor 1 (IGF1) is an example of an important stem cell communication molecule. If the activity of the growth factor is experimentally enhanced, muscle regeneration improves.

 

DNA Repair

DNA repair refers to a collection of processes by which a cell identifies and corrects damage to the DNA molecules that encode its genome. In human cells, both normal metabolic activities and environmental factors such as UV light can cause DNA damage, resulting in as many as 1 million individual molecular lesions per cell per day.Many of these lesions cause structural damage to the DNA molecule and can alter or eliminate the cell's ability to transcribe the gene that the affected DNA encodes. Other lesions induce potentially harmful mutations in the cell's genome, which affect the survival of its daughter cells after it undergoes mitosis. Consequently, the DNA repair process is constantly active as it responds to damage in the DNA structure.

The rate of DNA repair is dependent on many factors, including the cell type, the age of the cell, and the extracellular environment. A cell that has accumulated a large amount of DNA damage, or one that no longer effectively repairs damage incurred to its DNA, can enter one of three possible states:

1. an irreversible state of dormancy, known as senescence
2. cell suicide, also known as apoptosis or programmed cell death
3. unregulated cell division, which can lead to the formation of a tumor that is cancerous

The DNA repair ability of a cell is vital to the integrity of its genome and thus to its normal functioning and that of the organism. Many genes that were initially shown to influence lifespan have turned out to be involved in DNA damage repair and protection. Failure to correct molecular lesions in cells that form gametes can introduce mutations into the genomes of the offspring and thus influence the rate of evolution.

DNA damage

DNA damage, due to environmental factors and normal metabolic processes inside the cell, occurs at a rate of 1,000 to 1,000,000 molecular lesions per cell per day. While this constitutes only 0.000165% of the human genome's approximately 6 billion bases (3 billion base pairs), unrepaired lesions in critical genes (such as tumor suppressor genes) can impede a cell's ability to carry out its function and appreciably increase the likelihood of tumor formation.

The vast majority of DNA damage affects the primary structure of the double helix; that is, the bases themselves are chemically modified. These modifications can in turn disrupt the molecules' regular helical structure by introducing non-native chemical bonds or bulky adducts that do not fit in the standard double helix. Unlike proteins and RNA, DNA usually lacks tertiary structure and therefore damage or disturbance does not occur at that level. DNA is, however, supercoiled and wound around "packaging" proteins called histones (in eukaryotes), and both superstructures are vulnerable to the effects of DNA damage.

DNA repair mechanisms

Cells cannot function if DNA damage corrupts the integrity and accessibility of essential information in the genome (but cells remain superficially functional when so-called "non-essential" genes are missing or damaged). Depending on the type of damage inflicted on the DNA's double helical structure, a variety of repair strategies restore lost information. If possible, cells use the unmodified complementary strand of the DNA or the sister chromatid as a template to losslessly recover the original information. Without access to a template, cells use an error-prone recovery mechanism known as translesion synthesis as a last resort.

Damage to DNA alters the spatial configuration of the helix and such alterations can be detected by the cell. Once damage is localized, specific DNA repair molecules bind at or near the site of damage, inducing other molecules to bind and form a complex that enables the actual repair to take place. The types of molecules involved and the mechanism of repair that is mobilized depend on the type of damage that has occurred and the phase of the cell cycle that the cell is in.

Direct reversal

Cells are known to eliminate three types of damage to their DNA by chemically reversing it. These mechanisms do not require a template, since the types of damage they counteract can only occur in one of the four bases. Such direct reversal mechanisms are specific to the type of damage incurred and do not involve breakage of the phosphodiester backbone. The formation of thymine dimers (a common type of cyclobutyl dimer) upon irradiation with UV light results in an abnormal covalent bond between adjacent thymidine bases. The photoreactivation process directly reverses this damage by the action of the enzyme photolyase, whose activation is obligately dependent on energy absorbed from blue/UV light (300–500nm wavelength) to promote catalysis. Another type of damage, methylation of guanine bases, is directly reversed by the protein methyl guanine methyl transferase (MGMT), the bacterial equivalent of which is called as ogt. This is an expensive process because each MGMT molecule can only be used once; that is, the reaction is stoichiometric rather than catalytic.A generalized response to methylating agents in bacteria is known as the adaptive response and confers a level of resistance to alkylating agents upon sustained exposure by upregulation of alkylation repair enzymes. The third type of DNA damage reversed by cells is certain methylation of the bases cytosine and adenine.

When only one of the two strands of a double helix has a defect, the other strand can be used as a template to guide the correction of the damaged strand. In order to repair damage to one of the two paired molecules of DNA, there exist a number of excision repair mechanisms that remove the damaged nucleotide and replace it with an undamaged nucleotide complementary to that found in the undamaged DNA strand.

1. Base excision repair (BER), which repairs damage to a single nucleotide caused by oxidation, alkylation, hydrolysis, or deamination. The base is removed with glycosylase and ultimately replaced by repair synthesis with DNA ligase.
2. Nucleotide excision repair (NER), which repairs damage affecting longer strands of 2–30 bases. This process recognizes bulky, helix-distorting changes such as thymine dimers as well as single-strand breaks (repaired with enzymes such UvrABC endonuclease). A specialized form of NER known as Transcription-Coupled Repair (TCR) deploys high-priority NER repair enzymes to genes that are being actively transcribed.
3. Mismatch repair (MMR), which corrects errors of DNA replication and recombination that result in mispaired (but normal, that is non- damaged) nucleotides following DNA replication.
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Double-strand breaks

Double-strand breaks (DSBs), in which both strands in the double helix are severed, are particularly hazardous to the cell because they can lead to genome rearrangements. Two mechanisms exist to repair DSBs: non-homologous end joining (NHEJ) and recombinational repair

, a specialized DNA Ligase that forms a complex with the cofactor XRCC4, directly joins the two ends. To guide accurate repair, NHEJ relies on short homologous sequences called microhomologies present on the single-stranded tails of the DNA ends to be joined. If these overhangs are compatible, repair is usually accurate. NHEJ can also introduce mutations during repair. Loss of damaged nucleotides at the break site can lead to deletions, and joining of nonmatching termini forms translocations. NHEJ is especially important before the cell has replicated its DNA, since there is no template available for repair by homologous recombination. There are "backup" NHEJ pathways in higher eukaryotes. Besides its role as a genome caretaker, NHEJ is required for joining hairpin-capped double-strand breaks induced during V(D)J recombination, the process that generates diversity in B-cell and T-cell receptors in the vertebrate immune system.

Recombinational repair requires the presence of an identical or nearly identical sequence to be used as a template for repair of the break. The enzymatic machinery responsible for this repair process is nearly identical to the machinery responsible for chromosomal crossover during meiosis. This pathway allows a damaged chromosome to be repaired using a sister chromatid (available in G2 after DNA replication) or a homologous chromosome as a template. DSBs caused by the replication machinery attempting to synthesize across a single-strand break or unrepaired lesion cause collapse of the replication fork and are typically repaired by recombination.

Topoisomerases introduce both single- and double-strand breaks in the course of changing the DNA's state of supercoiling, which is especially common in regions near an open replication fork. Such breaks are not considered DNA damage because they are a natural intermediate in the topoisomerase biochemical mechanism and are immediately repaired by the enzymes that created them.

A team of French researchers bombarded Deinococcus radiodurans to study the mechanism of double-strand break DNA repair in that organism. At least two copies of the genome, with random DNA breaks, can form DNA fragments through annealing. Partially overlapping fragments are then used for synthesis of homologous regions through a moving D-loop that can continue extension until they find complementary partner strands. In the final step there is crossover by means of RecA-dependent homologous recombination.

Translesion synthesis

Translesion synthesis is a DNA damage tolerance process that allows the DNA replication machinery to replicate past DNA lesions such as thymine dimers or AP sites. It involves the switching out of regular DNA polymerases for specialized translesion polymerases, often with larger active sites that can facilitate the insertion of bases opposite damaged nucleotides. The polymerase switching is thought to be mediated by, among other factors, the post-translational modification of the replication processivity factor PCNA. Translesion synthesis polymerases often have low fidelity (high propensity to insert wrong bases) relative to regular polymerases. However, many are extremely efficient at inserting correct bases opposite specific types of damage. For example, Pol η mediates error-free bypass of lesions induced by UV irradiation, whereas Pol ζ introduces mutations at these sites. From a cellular perspective, risking the introduction of point mutations during translesion synthesis may be preferable to resorting to more drastic mechanisms of DNA repair, which may cause gross chromosomal aberrations or cell death.