BIOE 402. Medical Technology Assessment. 2 or 3 hours. Bioentrepreneur course. Assessment of medical technology in the context of commercialization. Objectives, competition, market share, funding, pricing, manufacturing, growth, and intellectual property; many issues unique to biomedical products. Course Information: 2 undergraduate hours. 3 graduate hours. Prerequisite(s): Junior standing or above and consent of the instructor.

“Bioinformatics” 특집을 내면서

Metabolic engineering of Clostridium autoethanogenum for selective alcohol production

Synthesis gas (syngas) is a gas mixture consisting mainly of H2, CO, and CO2 and can be derived from different sources, including renewable materials like lignocellulose. The fermentation of syngas to certain biofuels, using acetogenic bacteria, has attracted more and more interest over the last years. However, this technology is limited by two things: (1) the lack of complete knowledge of the energy metabolism of acetogenic bacteria, and (2) the lack of sophisticated genetic tools for the modification of acetogens. In this review, we discuss the bioenergetic constraints for the conversion of syngas to different biofuels. We will mainly focus on Acetobacterium woodii, which is the best understood acetogen in terms of energy conservation. Syngas fermentation with Clostridium autoethanogenum will also be discussed, since this organism is well suited to convert syngas to certain products and already used in large-scale industrial processes.

/pdf/bioenergetic-constraints-for-conversion-of-syngas-to-4f24cgk237.pdf

Bioenergetic constraints for conversion of syngas to biofuels in acetogenic bacteria

Ethanol production by engineered thermophiles

Ruminococcus albus 7 has played a key role in the development of the concept of interspecies hydrogen transfer. The rumen bacterium ferments glucose to 1.3 acetate, 0.7 ethanol, 2 CO2, and 2.6 H2 when growing in batch culture and to 2 acetate, 2 CO2, and 4 H2 when growing in continuous culture in syntrophic association with H2-consuming microorganisms that keep the H2 partial pressure low. The organism uses NAD+ and ferredoxin for glucose oxidation to acetyl coenzyme A (acetyl-CoA) and CO2, NADH for the reduction of acetyl-CoA to ethanol, and NADH and reduced ferredoxin for the reduction of protons to H2. Of all the enzymes involved, only the enzyme catalyzing the formation of H2 from NADH remained unknown. Here, we report that R. albus 7 grown in batch culture on glucose contained, besides a ferredoxin-dependent [FeFe]-hydrogenase (HydA2), a ferredoxin- and NAD-dependent electron-bifurcating [FeFe]-hydrogenase (HydABC) that couples the endergonic formation of H2 from NADH to the exergonic formation of H2 from reduced ferredoxin. Interestingly, hydA2 is adjacent to the hydS gene, which is predicted to encode an [FeFe]-hydrogenase with a C-terminal PAS domain. We showed that hydS and hydA2 are part of a larger transcriptional unit also harboring putative genes for a bifunctional acetaldehyde/ethanol dehydrogenase (Aad), serine/threonine protein kinase, serine/threonine protein phosphatase, and a redox-sensing transcriptional repressor. Since HydA2 and Aad are required only when R. albus grows at high H2 partial pressures, HydS could be a H2-sensing [FeFe]-hydrogenase involved in the regulation of their biosynthesis.

Hydrogen Formation and Its Regulation in Ruminococcus albus: Involvement of an Electron-Bifurcating [FeFe]-Hydrogenase, of a Non-Electron-Bifurcating [FeFe]-Hydrogenase, and of a Putative Hydrogen-Sensing [FeFe]-Hydrogenase

Bifunctional alcohol/aldehyde dehydrogenase (ADHE) enzymes are found within many fermentative microorganisms. They catalyse the conversion of an acyl-coenzyme A to an alcohol via an aldehyde intermediate; this is coupled to the oxidation of two NADH molecules to maintain the NAD+ pool during fermentative metabolism. The structure of the alcohol dehydrogenase (ADH) domain of an ADHE protein from the ethanol-producing thermophile Geobacillus thermoglucosidasius has been determined to 2.5 A resolution. This is the first structure to be reported for such a domain. In silico modelling has been carried out to generate a homology model of the aldehyde dehydrogenase domain, and this was subsequently docked with the ADH-domain structure to model the structure of the complete ADHE protein. This model suggests, for the first time, a structural mechanism for the formation of the large multimeric assemblies or `spirosomes' that are observed for this ADHE protein and which have previously been reported for ADHEs from other organisms.

/pdf/structure-of-a-bifunctional-alcohol-dehydrogenase-involved-byd4b76oiq.pdf

Structure of a bifunctional alcohol dehydrogenase involved in bioethanol generation in Geobacillus thermoglucosidasius

Acetylating aldehyde dehydrogenases (AcAldDH) catalyse the acetylation of Coenzyme-A (CoA), or in reverse generate acetaldehyde from Acetyl-CoA using NADH as a co-factor. This article reports the expression, purification, enzyme assay, and X-ray crystal structures of an AcAldDH from Geobacillus thermoglucosidasius (GtAcAldDH) to 2.1A and in complex with CoA and NAD+ to 4.0A. In the structure, the AcAldDH forms a close-knit dimer, similar to that seen in other Alcohol Dehydrogenase (ADH) structures. In GtAcAldDH, these dimers associate via their N-termini to form weakly interacting tetramers. This mode of tetrameric association is also seen in an unpublished AcAldDH deposited in the PDB, but is in contrast to all other ADH structures, (including the one other published AcAldDH found in a bacterial microcompartment), in which the dimers bury a large surface area including the C-termini. This novel mode of association sequesters the active sites and potentially reactive acyl-enzyme intermediates in the center of the tetramer. In other respects, the structure is very similar to the other AcAldDH, binding the cofactors in a corresponding fashion. This similarity enabled the identification of a shortened substrate cavity in G. thermoglucosidasius AcAldDH, explaining the limitations on the length of substrate accepted by the enzyme.

/pdf/structure-of-an-acetylating-aldehyde-dehydrogenase-from-the-4jwzzpo5gs.pdf

Jonathan Extance

Papers

Structure of a bifunctional alcohol dehydrogenase involved in bioethanol generation in Geobacillus thermoglucosidasius

Structure of an acetylating aldehyde dehydrogenase from the thermophilic ethanologen Geobacillus thermoglucosidasius.