https://web.iitd.ac.in/~sghosh08/publications/actabio2015.pdf

Silk fibroin as biomaterial for bone tissue engineering

Global climate change linked to the accumulation of greenhouse gases has caused concerns regarding the use of fossil fuels as the major energy source. To mitigate climate change while keeping energy supply sustainable, one solution is to rely on the ability of microorganisms to use renewable resources for biofuel synthesis. In this Review, we discuss how microorganisms can be explored for the production of next-generation biofuels, based on the ability of bacteria and fungi to use lignocellulose; through direct CO2 conversion by microalgae; using lithoautotrophs driven by solar electricity; or through the capacity of microorganisms to use methane generated from landfill. Furthermore, we discuss how to direct these substrates to the biosynthetic pathways of various fuel compounds and how to optimize biofuel production by engineering fuel pathways and central metabolism.

/pdf/fuelling-the-future-microbial-engineering-for-the-production-4zayx8tgii.pdf

Fuelling the future: microbial engineering for the production of sustainable biofuels

Solar-powered electrochemical reduction of CO2 and H2O to syngas, followed by fermentation, could lead to sustainable production of useful chemicals. However, due to insufficient electric current densities and instabilities of current CO2-to-CO electrolysers, a practical, scalable artificial photosynthesis remains a major challenge. Here, we address these problems using a commercially available silver-based gas diffusion electrode (used in industrial-scale chlorine–alkaline electrolysis) as the cathode in the CO2 electrolyser. Electric current densities up to 300 mA cm–2 were demonstrated for more than 1,200 hours with continuous operation. This CO2 electrolyser was coupled to a fermentation module, where the out-coming syngas from the CO2 electrolyser was converted to butanol and hexanol with high carbon selectivity. Conversion of photovoltaic electricity, CO2 and H2O to the desired alcohols achieved close to 100% Faradaic efficiency. Industrial production of useful and high-value chemicals via artificial photosynthesis is closer than expected with the proposed scalable hybrid system. The generation of useful chemicals from CO2 and renewable energy is an attractive—but challenging—endeavour. This work reports on the long-term operation of commercial electrodes for efficient CO2 reduction, with subsequent fermentation of the syngas product completing the technical photosynthesis of alcohols.

Technical photosynthesis involving CO 2 electrolysis and fermentation

Harnessing lipogenic pathways and rewiring acyl-CoA and acyl-ACP (acyl carrier protein) metabolism in Yarrowia lipolytica hold great potential for cost-efficient production of diesel, gasoline-like fuels, and oleochemicals. Here we assessed various pathway engineering strategies in Y. lipolytica toward developing a yeast biorefinery platform for sustainable production of fuel-like molecules and oleochemicals. Specifically, acyl-CoA/acyl-ACP processing enzymes were targeted to the cytoplasm, peroxisome, or endoplasmic reticulum to generate fatty acid ethyl esters and fatty alkanes with tailored chain length. Activation of endogenous free fatty acids and the subsequent reduction of fatty acyl-CoAs enabled the efficient synthesis of fatty alcohols. Engineering a hybrid fatty acid synthase shifted the free fatty acids to a medium chain-length scale. Manipulation of alternative cytosolic acetyl-CoA pathways partially decoupled lipogenesis from nitrogen starvation and unleashed the lipogenic potential of Y. lipolytica. Taken together, the strategies reported here represent promising steps to develop a yeast biorefinery platform that potentially upgrades low-value carbons to high-value fuels and oleochemicals in a sustainable and environmentally friendly manner.

https://www.pnas.org/content/pnas/113/39/10848.full.pdf

Engineering Yarrowia lipolytica as a platform for synthesis of drop-in transportation fuels and oleochemicals

There is an immediate need to drastically reduce the emissions associated with global fossil fuel consumption in order to limit climate change. However, carbon-based materials, chemicals, and transportation fuels are predominantly made from fossil sources and currently there is no alternative source available to adequately displace them. Gas-fermenting microorganisms that fix carbon dioxide (CO2) and carbon monoxide (CO) can break this dependence as they are capable of converting gaseous carbon to fuels and chemicals. As such, the technology can utilize a wide range of feedstocks including gasified organic matter of any sort (e.g., municipal solid waste, industrial waste, biomass, and agricultural waste residues) or industrial off-gases (e.g., from steel mills or processing plants). Gas fermentation has matured to the point that large-scale production of ethanol from gas has been demonstrated by two companies. This review gives an overview of the gas fermentation process, focusing specifically on anaerobic acetogens. Applications of synthetic biology and coupling gas fermentation to additional processes are discussed in detail. Both of these strategies, demonstrated at bench-scale, have abundant potential to rapidly expand the commercial product spectrum of gas fermentation and further improve efficiencies and yields.

/pdf/gas-fermentation-a-flexible-platform-for-commercial-scale-4m7a4ek8u8.pdf

Gas Fermentation—A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks

Engineering lipid overproduction in the oleaginous yeast Yarrowia lipolytica

In the quest for inexpensive feedstocks for the cost-effective production of liquid fuels, we have examined gaseous substrates that could be made available at low cost and sufficiently large scale for industrial fuel production. Here we introduce a new bioconversion scheme that effectively converts syngas, generated from gasification of coal, natural gas, or biomass, into lipids that can be used for biodiesel production. We present an integrated conversion method comprising a two-stage system. In the first stage, an anaerobic bioreactor converts mixtures of gases of CO2 and CO or H2 to acetic acid, using the anaerobic acetogen Moorella thermoacetica The acetic acid product is fed as a substrate to a second bioreactor, where it is converted aerobically into lipids by an engineered oleaginous yeast, Yarrowia lipolytica We first describe the process carried out in each reactor and then present an integrated system that produces microbial oil, using synthesis gas as input. The integrated continuous bench-scale reactor system produced 18 g/L of C16-C18 triacylglycerides directly from synthesis gas, with an overall productivity of 0.19 g⋅L(-1)⋅h(-1) and a lipid content of 36%. Although suboptimal relative to the performance of the individual reactor components, the presented integrated system demonstrates the feasibility of substantial net fixation of carbon dioxide and conversion of gaseous feedstocks to lipids for biodiesel production. The system can be further optimized to approach the performance of its individual units so that it can be used for the economical conversion of waste gases from steel mills to valuable liquid fuels for transportation.

https://www.pnas.org/content/pnas/113/14/3773.full.pdf

Integrated bioprocess for conversion of gaseous substrates to liquids.

Freeze-gelled silk fibroin protein scaffolds for potential applications in soft tissue engineering

Some aspects of this invention provide engineered microbes for oil production. Methods for microbe engineering and for use of engineered microbes are also provided herein. In some embodiments, microbes are provided that are engineered to modulate a combination of rate-controlling steps of lipid synthesis, for example, a combination of a step generating metabolites, acetyl-CoA, ATP or NADPH for lipid synthesis (a push step), and a step sequestering a product or an intermediate of a lipid synthesis pathway that mediates feedback inhibition of lipid synthesis (a pull step). Such push-and-pull engineered microbes exhibit greatly enhanced conversion yields and TAG synthesis and storage properties.

Engineered microbes and methods for microbial oil production

Abstract Annotating protein sequences according to their biological functions is one of the key steps in understanding microbial diversity, metabolic potentials, and evolutionary histories. However, even in the best-studied prokaryotic genomes, not all proteins can be characterized by classical in vivo, in vitro, and/or in silico methods—a challenge rapidly growing alongside the advent of next-generation sequencing technologies and their enormous extension of ‘omics’ data in public databases. These so-called hypothetical proteins (HPs) represent a huge knowledge gap and hidden potential for biotechnological applications. Opportunities for leveraging the available ‘Big Data’ have recently proliferated with the use of artificial intelligence (AI). Here, we review the aims and methods of protein annotation and explain the different principles behind machine and deep learning algorithms including recent research examples, in order to assist both biologists wishing to apply AI tools in developing comprehensive genome annotations and computer scientists who want to contribute to this leading edge of biological research.

/pdf/elucidating-the-functional-roles-of-prokaryotic-proteins-1jf2bv1g.pdf

Sagar Chakraborty

Papers

Engineering lipid overproduction in the oleaginous yeast Yarrowia lipolytica

Integrated bioprocess for conversion of gaseous substrates to liquids.

Freeze-gelled silk fibroin protein scaffolds for potential applications in soft tissue engineering

Engineered microbes and methods for microbial oil production

Elucidating the functional roles of prokaryotic proteins using big data and artificial intelligence