Electron transport chain in aerobically cultivated Zymomonas mobilis

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ENERGY COUPLING SITES IN THE ELECTRON-TRANSPORT CHAIN OF ZYMOMONAS-MOBILIS

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Effect of ADH II Deficiency on the Intracellular Redox Homeostasis in Zymomonas mobilis

Mutant strain of the facultatively anaerobic, ethanol-producing bacterium Zymomonas mobilis, deficient in the Fe-containing alcohol dehydrogenase isoenzyme (ADH II), showed impaired homeostasis of the intracellular NAD(P)H during transition from anaerobic to aerobic conditions, and also in steady-state continuous cultures at various oxygen supplies. At the same time, ADH II deficiency in aerobically grown cells was accompanied by a threefold increase of catalase activity and by about 50% increase of hydrogen peroxide excretion. It is concluded that ADH II under aerobic conditions functions to maintain intracellular redox homeostasis and to protect the cells from endogenous hydrogen peroxide

Crypthecodinium cohnii and Zymomonas mobilis syntrophy for production of omega 3 fatty acid

One of missions of industrial biotechnology is to produce valuable products from waste. In the current study we aim to establish a bioprocess of syntrophic co-cultivation of two microorganisms (Crypthecodinium cohnii and Zymomonas mobilis) at laboratory prototype level for bioconversion of renewables (molasses, the byproduct of sugar industry, and glycerol, byproduct of biodiesel production) into valuable vegetarian (produced by microorganisms) docosahexaenoic acid (DHA) from omega 3 fatty acid family. Cells of Z.mobilis strain (respiratory-deficient mutant derivative of the wild type strain Z.mobilis ATCC 29191) will be immobilized, following the previously described procedure of calcium alginate gel encapsulation. Mathematical modelling will be used for different components and at different stages of the syntrophic process to assist its improvement. The syntrophic bioprocess will be optimized with respect to (i) the medium composition, (ii) feed rates, (iii) oxygen supply, and (iv) the amounts of added immobilized Z. mobilis cells at particular stages of the fermentation. Genome and/or central carbon metabolism scale stoichiometric modelling and optimization will be performed to increase the utilization of cheap substrates and pre-treated residuals of C.cohnii. Bioprocess dynamics with respect to growth speed of batch fermentations will be modelled to ensure optimal proportions of syntrophic organisms, taking into account Z.mobilis production and immobilization as “slave” process of C.cohnii cultivation. Within several iterative cycles the fermentation data will be used for modelling and computer simulation of the bioprocess, and at the same time, the model-predicted dynamic behaviour of the system will aid further improvement of fermentation performance.publishersversionPeer reviewe

Modeling of Zymomonas mobilis central metabolism for novel metabolic engineering strategies

Mathematical modeling of metabolism is essential for rational metabolic engineering. The present work focuses on several types of modeling approach to quantitative understanding of central metabolic network and energetics in the bioethanol-producing bacterium Zymomonas mobilis. Combined use of Flux Balance, Elementary Flux Mode, and thermodynamic analysis of its central metabolism, together with dynamic modeling of the core catabolic pathways, can help to design novel substrate and product pathways by systematically analyzing the solution space for metabolic engineering, and yields insights into the function of metabolic network, hardly achievable without applying modeling tools

Application of FT-IR Spectroscopy for Fingerprinting of Zymomonas mobilis Respiratory Mutants

Abstract. Z. mobilis ATCC 29191 and its respiratory knockout mutants, kat-, ndh-, cytB-, and cydB-, were grown under anaerobic and aerobic conditions. FT-IR spectroscopy was used to study the variations of the cell macromolecular composition. Quantitative analysis showed that the concentration ratios-nucleic acids to lipids, for Z. mobilis parent strain, kat-, ndh-, cytB-, and cydB-strains, clearly distinguished Z. mobilis parent strain from its mutant derivatives and corresponded fairly well to the expected degree of biochemical similarity between the strains. Two different FT-IR-spectra hierarchical cluster analysis (HCA) methods were created to differentiate Z. mobilis parent strain and respiratory knockout mutant strains. HCA based on discriminative spectra ranges of carbohydrates, nucleic acids, and lipids allowed to evaluate the influence of growth environment (aeration, growth phase) on the macromolecular composition of cells and differentiate the strains. HCA based on IR spectra of inoculums, in a diagnostic region including the characteristic nucleic acid vibration modes, clearly discriminated the strains under study. Thus it was shown that FT-IR spectroscopy can distinguish various alterations of Z. mobilis respiratory metabolism by HCA of biomass spectra

Structure and function of the bacterial heterodimeric ABC transporter CydDC: stimulation of ATPase activity by thiol and heme compounds.

In Escherichia coli, the biogenesis of both cytochrome bd-type quinol oxidases and periplasmic cytochromes requires the ATP-binding cassette-type cysteine/GSH transporter, CydDC. Recombinant CydDC was purified as a heterodimer and found to be an active ATPase both in soluble form with detergent and when reconstituted into a lipid environment. Two-dimensional crystals of CydDC were analyzed by electron cryomicroscopy, and the protein was shown to be made up of two non-identical domains corresponding to the putative CydD and CydC subunits, with dimensions characteristic of other ATP-binding cassette transporters. CydDC binds heme b. Detergent-solubilized CydDC appears to adopt at least two structural states, each associated with a characteristic level of bound heme. The purified protein in detergent showed a weak basal ATPase activity (approximately 100 nmol Pi/min/mg) that was stimulated ∼3-fold by various thiol compounds, suggesting that CydDC could act as a thiol transporter. The presence of heme (either intrinsic or added in the form of hemin) led to a further enhancement of thiol-stimulated ATPase activity, although a large excess of heme inhibited activity. Similar responses of the ATPase activity were observed with CydDC reconstituted into E. coli lipids. These results suggest that heme may have a regulatory role in CydDC-mediated transmembrane thiol transport

Transcriptomic and metabolomic profiling of Zymomonas mobilis during aerobic and anaerobic fermentations

<p>Abstract</p> <p>Background</p> <p><it>Zymomonas mobilis </it>ZM4 (ZM4) produces near theoretical yields of ethanol with high specific productivity and recombinant strains are able to ferment both C-5 and C-6 sugars. <it>Z. mobilis </it>performs best under anaerobic conditions, but is an aerotolerant organism. However, the genetic and physiological basis of ZM4's response to various stresses is understood poorly.</p> <p>Results</p> <p>In this study, transcriptomic and metabolomic profiles for ZM4 aerobic and anaerobic fermentations were elucidated by microarray analysis and by high-performance liquid chromatography (HPLC), gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS) analyses. In the absence of oxygen, ZM4 consumed glucose more rapidly, had a higher growth rate, and ethanol was the major end-product. Greater amounts of other end-products such as acetate, lactate, and acetoin were detected under aerobic conditions and at 26 h there was only 1.7% of the amount of ethanol present aerobically as there was anaerobically. In the early exponential growth phase, significant differences in gene expression were not observed between aerobic and anaerobic conditions via microarray analysis. HPLC and GC analyses revealed minor differences in extracellular metabolite profiles at the corresponding early exponential phase time point.</p> <p>Differences in extracellular metabolite profiles between conditions became greater as the fermentations progressed. GC-MS analysis of stationary phase intracellular metabolites indicated that ZM4 contained lower levels of amino acids such as alanine, valine and lysine, and other metabolites like lactate, ribitol, and 4-hydroxybutanoate under anaerobic conditions relative to aerobic conditions. Stationary phase microarray analysis revealed that 166 genes were significantly differentially expressed by more than two-fold. Transcripts for Entner-Doudoroff (ED) pathway genes (<it>glk, zwf, pgl, pgk, and eno</it>) and gene <it>pdc</it>, encoding a key enzyme leading to ethanol production, were at least 30-fold more abundant under anaerobic conditions in the stationary phase based on quantitative-PCR results. We also identified differentially expressed ZM4 genes predicted by The Institute for Genomic Research (TIGR) that were not predicted in the primary annotation.</p> <p>Conclusion</p> <p>High oxygen concentrations present during <it>Z. mobilis </it>fermentations negatively influence fermentation performance. The maximum specific growth rates were not dramatically different between aerobic and anaerobic conditions, yet oxygen did affect the physiology of the cells leading to the buildup of metabolic byproducts that ultimately led to greater differences in transcriptomic profiles in stationary phase.</p