Transgenic E. coli made to produce hydrocarbon fuels

Super cool work from researchers in the UK

Excerpt from “Bacteria churn out first ever petrol-like biofuel” by Rebecca Summers

To be used as a mainstream alternative to fossil fuels – desirable because biofuels are carbon-neutral over their lifetime – engines would have to be redesigned, or an extra processing step employed to convert the fuel into a more usable form.

To try to bypass that, John Love from the University of Exeter in the UK and colleagues took genes from the camphor tree, soil bacteria and blue-green algae and spliced them into DNA from Escherichia coli bacteria. When the modified E. coli were fed glucose, the enzymes they produced converted the sugar into fatty acids and then turned these into hydrocarbons that were chemically and structurally identical to those found in commercial fuel.

“We are biologically producing the fuel that the oil industry makes and sells,” says Love.

“Synthesis of customized petroleum-replica fuel molecules by targeted modification of free fatty acid pools in Escherichia coli”

Biofuels are the most immediate, practical solution for mitigating dependence on fossil hydrocarbons, but current biofuels (alcohols and biodiesels) require significant downstream processing and are not fully compatible with modern, mass-market internal combustion engines. Rather, the ideal biofuels are structurally and chemically identical to the fossil fuels they seek to replace (i.e., aliphatic n- and iso-alkanes and -alkenes of various chain lengths). Here we report on production of such petroleum-replica hydrocarbons in Escherichia coli. The activity of the fatty acid (FA) reductase complex from Photorhabdus luminescens was coupled with aldehyde decarbonylase from Nostoc punctiforme to use free FAs as substrates for alkane biosynthesis. This combination of genes enabled rational alterations to hydrocarbon chain length (Cn) and the production of branched alkanes through upstream genetic and exogenous manipulations of the FA pool. Genetic components for targeted manipulation of the FA pool included expression of a thioesterase from Cinnamomum camphora (camphor) to alter alkane Cn and expression of the branched-chain α-keto acid dehydrogenase complex and β-keto acyl-acyl carrier protein synthase III from Bacillus subtilis to synthesize branched (iso-) alkanes. Rather than simply reconstituting existing metabolic routes to alkane production found in nature, these results demonstrate the ability to design and implement artificial molecular pathways for the production of renewable, industrially relevant fuel molecules.

 

 

Researchers at GIT show spontaneous assembly of “proto-RNAs” in water

Really interesting work. Essentially shows that stacking interactions and hydrogen bonding of the nitrogenous bases is sufficient to form a “proto-RNA” filament. While far from being definitive proof of anything, this work certainly builds on and supports the RNA-world hypothesis for the origins of life. Very cool stuff, will update this post when more information becomes available.

Excerpt from “Molecules assemble in water, hint at origins of life” in e! Science News

 

Researchers at the Georgia Institute of Technology are exploring an alternate theory for the origin of RNA: they think the RNA bases may have evolved from a pair of molecules distinct from the bases we have today. This theory looks increasingly attractive, as the Georgia Tech group was able to achieve efficient, highly ordered self-assembly in water with small molecules that are similar to the bases of RNA. These “proto-RNA bases” spontaneously assemble into gene-length linear stacks, suggesting that the genes of life could have gotten started from these or similar molecules.

…

The discovery was made by a team of scientists led by Georgia Tech Professor Nicholas Hud, who has been trying for years to find simple molecules that will assemble in water and be capable of forming RNA or its ancestor. Hud’s group knew that they were on to something when they added a small chemical tail to a proto-RNA base and saw it spontaneously form linear assemblies with another proto-RNA base. In some cases, the results produced 18,000 nicely ordered, stacked molecules in one long structure.

…

Hud concedes that scientists may never be 100 percent sure what existed four billion years ago when a complex mixture of chemicals started to work together to start life. His next goal is to determine whether the proto-RNA bases can be linked by a backbone to form a polymer that could have functioned as a genetic material.

 

Some budding yeast I used to grow

Gotye Parody, hilarious.

Chemistry Dubstep

Amazing microbial diversity uncovered in the “Jungle” of our belly buttons

In a bizarre, but very interesting (and potentially hilarious) bit of science, Robert Dunn’s group at UNCS started investigating the “microbiome” of the human belly button. Using 16S rDNA libraries they analyzed 60 belly buttons they isolated 2,368 bacterial species, of which 1,458 could be new to science. Most people appeared to have about 67 species inhabiting their belly buttons. While the initial sample size was quite small it was still a stunning result to find so much microbial diversity inhabiting our oft forgotten belly buttons.

From “Microbial fauna in your belly button is like a ‘tropical forest” by Mihai Andrei

The whole thing started about two years ago, when an undergrad sampled a colleague’s belly button bacteria to send it to him as a Christmas card. Biologists, the quirky people that they are, quickly picked up on this idea.

The basic idea is that the belly button is one of the least scrubbed places of the human body, making it one of the most pristine bacterial environments humans harbor – which could kind of explain why some people are totally grossed out by navels.

….

Not even a single strain appeared on all subjects, but 8 were found in over 70 percent of subjects; and when one of the species found often was present, others followed in great numbers.

“That makes the belly button a lot like rain forests,” Dunn said. In any given forest, he explained, the spectrum of flora might vary, but an ecologist can count on a certain few dominant tree types. “The idea that some aspects of our bodies are like a rain forest—to me it’s quite beautiful,” he added. “And it makes sense to me as an ecologist. I understand what steps to take next; I can see how that works.”

The study was published in PLOS ONE: A Jungle in There: Bacteria in Belly Buttons are Highly Diverse, but Predictable

The belly button is one of the habitats closest to us, and yet it remains relatively unexplored. We analyzed bacteria and arachaea from the belly buttons of humans from two different populations sampled within a nation-wide citizen science project. We examined bacterial and archaeal phylotypes present and their diversity using multiplex pyrosequencing of 16S rDNA libraries. We then tested the oligarchy hypothesis borrowed from tropical macroecology, namely that the frequency of phylotypes in one sample of humans predicts its frequency in another independent sample. We also tested the predictions that frequent phylotypes (the oligarchs) tend to be common when present, and tend to be more phylogenetically clustered than rare phylotypes. Once rarefied to four hundred reads per sample, bacterial communities from belly buttons proved to be at least as diverse as communities known from other skin studies (on average 67 bacterial phylotypes per belly button). However, the belly button communities were strongly dominated by a few taxa: only 6 phylotypes occurred on >80% humans. While these frequent bacterial phylotypes (the archaea were all rare) are a tiny part of the total diversity of bacteria in human navels (<0.3% of phylotypes), they constitute a major portion of individual reads (~1/3), and are predictable among independent samples of humans, in terms of both the occurrence and evolutionary relatedness (more closely related than randomly drawn equal sets of phylotypes). Thus, the hypothesis that “oligarchs” dominate diverse assemblages appears to be supported by human-associated bacteria. Although it remains difficult to predict which species of bacteria might be found on a particular human, predicting which species are most frequent (or rare) seems more straightforward, at least for those species living in belly buttons.

 

Unfortunate news for fair skinned folks, inherently higher chance of developing melanoma independent of sun exposure

Excerpt from “For redheads, skin cancer may be in the genes” by Kate Shaw

It turns out that redheads may be vulnerable to more oxidative DNA damage, even if they stay out of the sun: the researchers found several indicators of DNA damage—lipid peroxide and two cyclopurine levels—were much higher in mice that produced large amounts of pheomelanin.  This DNA damage, in turn, contributes to the development of malignant melanoma.

It has long been known that UV radiation causes skin cancer, especially in fair-skinned redheads, but these results suggest that even redheads that stay out of the sun aren’t completely protected from melanoma. The same pigment pathway that gives them their fiery hair and freckles may actually damage their DNA and lead to melanoma, without any help from the sun’s rays. Unfortunately, it’s still unclear how to stop this damage. Experts advise redheads to visit the dermatologist regularly, and suggest that increasing oral antioxidant intake might decrease melanoma risk.

Nature research article behind these finding “An ultraviolet-radiation-independent pathway to melanoma carcinogenesis in the red hair/fair skin background“

People with pale skin, red hair, freckles and an inability to tan—the ‘red hair/fair skin’ phenotype—are at highest risk of developing melanoma, compared to all other pigmentation types¹. Genetically, this phenotype is frequently the product of inactivating polymorphisms in the melanocortin 1 receptor (MC1R) gene. MC1R encodes a cyclic AMP-stimulating G-protein-coupled receptor that controls pigment production. Minimal receptor activity, as in red hair/fair skin polymorphisms, produces the red/yellow pheomelanin pigment, whereas increasing MC1R activity stimulates the production of black/brown eumelanin². Pheomelanin has weak shielding capacity against ultraviolet radiation relative to eumelanin, and has been shown to amplify ultraviolet-A-induced reactive oxygen species^{3, 4, 5}. Several observations, however, complicate the assumption that melanoma risk is completely ultraviolet-radiation-dependent. For example, unlike non-melanoma skin cancers, melanoma is not restricted to sun-exposed skin and ultraviolet radiation signature mutations are infrequently oncogenic drivers⁶. Although linkage of melanoma risk to ultraviolet radiation exposure is beyond doubt, ultraviolet-radiation-independent events are likely to have a significant role^{1, 7}. Here we introduce a conditional, melanocyte-targeted allele of the most common melanoma oncoprotein, BRAF^V600E, into mice carrying an inactivating mutation in the Mc1r gene (these mice have a phenotype analogous to red hair/fair skin humans). We observed a high incidence of invasive melanomas without providing additional gene aberrations or ultraviolet radiation exposure. To investigate the mechanism of ultraviolet-radiation-independent carcinogenesis, we introduced an albino allele, which ablates all pigment production on the Mc1r^e/e background. Selective absence of pheomelanin synthesis was protective against melanoma development. In addition, normal Mc1r^e/e mouse skin was found to have significantly greater oxidative DNA and lipid damage than albino-Mc1r^e/e mouse skin. These data suggest that the pheomelanin pigment pathway produces ultraviolet-radiation-independent carcinogenic contributions to melanomagenesis by a mechanism of oxidative damage. Although protection from ultraviolet radiation remains important, additional strategies may be required for optimal melanoma prevention.

a, C57BL/6 pigmentation variants with epidermal melanocytes (K14-SCF). From left to right: black (wild type), red (Mc1re/e) and albino (Tyrc/c). b, Genotype of animals used for experimental studies. c, Percentage survival of pigmentation variants not carrying the K14-SCF transgene, that is, no epidermal melanocytes (nblack = 28, nred = 40, nalbino = 48). Pblack–albino = 0.250, Pblack–red = 0.003, Palbino–red = 0.003. d, Percentage survival of pigmentation variants carrying the K14–SCF transgene, that is, epidermal melanocytes (nblack = 49, nred = 77, nalbino = 41). Pblack–albino = 0.103, Pblack–red = 0.009, Palbino–red < 0.0001.

 

New quick cook “method” for turns algae into biofuel at astonishing efficiency

This is really cool, pretty amazing they can get such high conversion rates. Might make algae based biofuels a reality.

Snippets from: Biofuel breakthrough: Quick cook method turns algae into oil

Michigan Engineering researchers can “pressure-cook” algae for as little as a minute and transform an unprecedented 65 percent of the green slime into biocrude.

“We’re trying to mimic the process in nature that forms crude oil with marine organisms,” said Phil Savage, an Arthur F. Thurnau professor and a professor of chemical engineering at the University of Michigan.

…..

One of the advantages of the wet method is that it doesn’t just extract the existing fat from the algae—it also breaks down proteins and carbohydrates. The minute method did this so successfully that the oil contained about 90 percent of the energy in the original algae.

“That result is near the upper bound of what is possible,” Savage said.

The abstract for “The Effects of Heating Rate and Reaction Time On Hydrothermal Liquefaction of Microalgae”

Julia L. Faeth and Phillip E. Savage, Chemical Engineering, University of Michigan, Ann Arbor, MI

Hydrothermal liquefaction (HTL) is one of many methods for biomass conversion to biofuels or biofuel precursors. HTL avoids energy-intensive drying steps, and is therefore more energy efficient for biomass with very high moisture content, like microalgae. The effects of many different parameters (including biomass concentration, nominal reaction temperature, catalysis, and reaction time) on product yields for HTL of microalgae have been reported in the literature.  However, a majority of the reaction times reported in HTL literature have been defined to ignore the time necessary to reach the nominal reaction temperature. Furthermore, the heating times reported vary greatly, from several minutes to a few hours.The time necessary to heat an HTL reaction mixture to the nominal reaction temperature is dependent on the heating rate. The heating rate of a reaction mixture has been observed to influence the product yields for HTL of woody biomass. Product yields for supercritical water gasification of glucose and cabbage are also influenced by heating rate. This presentation explores the influence of heating rate on the product yields for HTL of microalga Nannochloropsis sp. The influence of reaction time, including time necessary for the reaction mixture to reach the nominal reaction temperature, will also be discussed. These effects need to be considered during the development of an efficient HTL flow reaction process.

 

New “prime and pull” vaccine model offers new hope against retroviruses

Excerpt from ScienceTechDaily

New Model for Vaccination Against Genital Herpes

Until now, most efforts to develop a vaccine have focused on the immune system’s antibodies, or T cells, circulating through the body. When T cells encounter foreign invaders such as bacteria or viruses, they learn to recognize them and mount ever-stronger immune responses to fight them. But efforts to harness these circulating T cells have not been effective in organs such as the vagina, intestines, lung airways, and central nervous system, which restrict the entry of these “memory” T cells.

To investigate an alternative approach, the Yale team focused instead on peripheral tissue in the female genital tract, where viral exposure occurs. The challenge was to recruit virus-specific T cells into the vaginal mucosa without triggering a potentially harmful inflammatory response of the immune system.

Working with mice, they explored a two-part vaccine strategy they call “prime and pull.” The “priming” involved conventional vaccination to elicit a system-wide T cell response. The “pulling” involved recruitment of activated T cells directly into the vaginal tissue, via topical application, of chemokines — substances that help mobilize the immune cells.

 

The original study can be found here

A vaccine strategy that protects against genital herpes by establishing local memory T cells by Haina Shin & Akiko Iwasaki

Most successful existing vaccines rely on neutralizing antibodies, which may not require specific anatomical localization of B cells. However, efficacious vaccines that rely on T cells for protection have been difficult to develop, as robust systemic memory T-cell responses do not necessarily correlate with host protection¹. In peripheral sites, tissue-resident memory T cells provide superior protection compared to circulating memory T cells^{2, 3}. Here we describe a simple and non-inflammatory vaccine strategy that enables the establishment of a protective memory T-cell pool within peripheral tissue. The female genital tract, which is a portal of entry for sexually transmitted infections, is an immunologically restrictive tissue that prevents entry of activated T cells in the absence of inflammation or infection⁴. To overcome this obstacle, we developed a vaccine strategy that we term ‘prime and pull’ to establish local tissue-resident memory T cells at a site of potential viral exposure. This approach relies on two steps: conventional parenteral vaccination to elicit systemic T-cell responses (prime), followed by recruitment of activated T cells by means of topical chemokine application to the restrictive genital tract (pull), where such T cells establish a long-term niche and mediate protective immunity. In mice, prime and pull protocol reduces the spread of infectious herpes simplex virus 2 into the sensory neurons and prevents development of clinical disease. These results reveal a promising vaccination strategy against herpes simplex virus 2, and potentially against other sexually transmitted infections such as human immunodeficiency virus.

 

For the love of food… and microscopy

Terra Cibus: Food Photographed with A Scanning Electron Microscope by Caren Alpert

San Francisco-based fine art and commercial photographer Caren Alpert combines her loves for photography, food, and art in these gorgeous photos taken with an electron microscope. Alpert captures the microscopic, almost other-worldly surfaces of common foods such as Oreo cookies, shrimp, leaves, and candy, turning what might normally be a scientific endeavor into fine art. As amazing as the images look here I’ve linked each through to the high resolution version on her website so you can see them in greater detail. Alpert has upcoming shows at Bertha V.B. Lederer Gallery starting October 2, as well as a show called The Beauty + Biology of our Food at the Citigroup Center starting November 2. She also has limited edition prints for sale and you can find out more by contacting her here.

See the entire Terra Cibus gallery here

terra cibus no.34 / pop tart (450x magnification)

terra cibus no.23 / purple onion (230x magnification)

terra cibus no.3 / celery Leaf (85x magnification)

New type of vaccine works to inhibit pathogenicity of drug resistant Gram-negative bacilli

A collaboration of researchers published a paper today in mBio where they show that inhibiting the synthesis of lipopolysaccharides correlated with virulence was an effective “antibiotic” against Acinetobacter baumannii – a often deadly bacteria especially prevalent in hospitals. This research is really exciting because the antibiotic is a small molecular inhibitor known as LpxC which affects lipid A biosynthesis, but is not aimed at killing the bacteria – simply reducing it’s pathogenicity. With the number of effective antibiotics dwindling at an alarming rate, this is exceedingly good news for the medical community struggling with the challenges of “super-bugs”.

 

Excerpt from: New antibiotic cures disease by disarming pathogens, not killing them

“We found that strains that caused the rapidly lethal infections shed lipopolysaccharide [also called LPS or endotoxin] while growing. The more endotoxin shed, the more virulent the strain was,” says Spellberg. This pinpointed a new therapy target for the researchers: the endotoxin these bacteria shed in the body.

Blocking the synthesis of the endotoxin with a small molecule called LpxC-1 prevented infected mice from getting sick. Unlike traditional antibiotics, Spellberg says, LpxC-1 doesn’t kill the bacteria, it just shuts down the manufacture of the endotoxin and stops the body from mounting the inflammatory immune response to it that is the actual cause of death in seriously ill patients.

Spellberg says this is a direction few researchers have taken when exploring ways to treat infections but that it could make the difference in finding an effective drug. The results also highlight how important it is to find new, physiologically relevant ways of screening potential antibiotics for pathogens with a high degree of resistance, write the authors. Molecules like LpxC-1 that inhibit rather than kill bacteria wouldn’t pass muster with traditional antibiotic screens that are based on killing effectiveness.

 

The actual article from mBio: Inhibition of LpxC Protects Mice from Resistant Acinetobacter baumannii by Modulating Inflammation and Enhancing Phagocytosis

ABSTRACT

New treatments are needed for extensively drug-resistant (XDR) Gram-negative bacilli (GNB), such as Acinetobacter baumannii. Toll-like receptor 4 (TLR4) was previously reported to enhance bacterial clearance of GNB, including A. baumannii. However, here we have shown that 100% of wild-type mice versus 0% of TLR4-deficient mice died of septic shock due to A. baumannii infection, despite having similar tissue bacterial burdens. The strain lipopolysaccharide (LPS) content and TLR4 activation by extracted LPS did not correlate with in vivo virulence, nor did colistin resistance due to LPS phosphoethanolamine modification. However, more-virulent strains shed more LPS during growth than less-virulent strains, resulting in enhanced TLR4 activation. Due to the role of LPS in A. baumannii virulence, an LpxC inhibitor (which affects lipid A biosynthesis) antibiotic was tested. The LpxC inhibitor did not inhibit growth of the bacterium (MIC > 512 µg/ml) but suppressed A. baumannii LPS-mediated activation of TLR4. Treatment of infected mice with the LpxC inhibitor enhanced clearance of the bacteria by enhancing opsonophagocytic killing, reduced serum LPS concentrations and inflammation, and completely protected the mice from lethal infection. These results identify a previously unappreciated potential for the new class of LpxC inhibitor antibiotics to treat XDR A. baumannii infections. Furthermore, they have far-reaching implications for pathogenesis and treatment of infections caused by GNB and for the discovery of novel antibiotics not detected by standard in vitro screens.

IMPORTANCE Novel treatments are needed for infections caused by Acinetobacter baumannii, a Gram-negative bacterium that is extremely antibiotic resistant. The current study was undertaken to understand the immunopathogenesis of these infections, as a basis for defining novel treatments. The primary strain characteristic that differentiated virulent from less-virulent strains was shedding of Gram-negative lipopolysaccharide (LPS) during growth. A novel class of antibiotics, called LpxC inhibitors, block LPS synthesis, but these drugs do not demonstrate the ability to kill A. baumannii in vitro. We found that an LpxC inhibitor blocked the ability of bacteria to activate the sepsis cascade, enhanced opsonophagocytic killing of the bacteria, and protected mice from lethal infection. Thus, an entire new class of antibiotics which is already in development has heretofore-unrecognized potential to treat A. baumannii infections. Furthermore, standard antibiotic screens based on in vitro killing failed to detect this treatment potential of LpxC inhibitors for A. baumannii infections.