Dataset Information

Protein Acetylation and Butyrylation Regulate the Phenotype and Metabolic Shifts of the Endospore-forming Clostridium acetobutylicum.

ABSTRACT: Clostridium acetobutylicum is a strict anaerobic, endospore-forming bacterium, which is used for the production of the high energy biofuel butanol in metabolic engineering. The life cycle of C. acetobutylicum can be divided into two phases, with acetic and butyric acids being produced in the exponential phase (acidogenesis) and butanol formed in the stationary phase (solventogenesis). During the transitional phase from acidogenesis to solventogenesis and latter stationary phase, concentration peaks of the metabolic intermediates butyryl phosphate and acetyl phosphate are observed. As an acyl group donor, acyl-phosphate chemically acylates protein substrates. However, the regulatory mechanism of lysine acetylation and butyrylation involved in the phenotype and solventogenesis of C. acetobutylicum remains unknown. In our study, we conducted quantitative analysis of protein acetylome and butyrylome to explore the dynamic change of lysine acetylation and butyrylation in the exponential phase, transitional phase, and stationary phase of C. acetobutylicum Total 458 lysine acetylation sites and 1078 lysine butyrylation sites were identified in 254 and 373 substrates, respectively. Bioinformatics analysis uncovered the similarities and differences between the two acylation modifications in C. acetobutylicum Mutation analysis of butyrate kinase and the central transcriptional factor Spo0A was performed to characterize the unique role of lysine butyrylation in the metabolic pathway and sporulation process of C. acetobutylicum Moreover, quantitative proteomic assays were performed to reveal the relationship between protein features (e.g. gene expression level and lysine acylation level) and metabolites in the three growth stages. This study expanded our knowledge of lysine acetylation and butyrylation in Clostridia and constituted a resource for functional studies on lysine acylation in bacteria.

SUBMITTER: Xu JY

PROVIDER: S-EPMC5986239 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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Protein Acetylation and Butyrylation Regulate the Phenotype and Metabolic Shifts of the Endospore-forming Clostridium acetobutylicum.

Xu Jun-Yu JY Xu Zhen Z Liu XinXin X Tan Minjia M Ye Bang-Ce BC

Molecular & cellular proteomics : MCP 20180309 6

Clostridium acetobutylicum is a strict anaerobic, endospore-forming bacterium, which is used for the production of the high energy biofuel butanol in metabolic engineering. The life cycle of C. acetobutylicum can be divided into two phases, with acetic and butyric acids being produced in the exponential phase (acidogenesis) and butanol formed in the stationary phase (solventogenesis). During the transitional phase from acidogenesis to solventogenesis and latter stationary phase, co ...[more]

PMID: 29523768

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Project description:Butanol is an important industrial solvent and advanced biofuel that can be produced by biphasic fermentation by Clostridium acetobutylicum. It has been known that acetate and butyrate first formed during the acidogenic phase are reassimilated to form acetone-butanol-ethanol (cold channel). Butanol can also be formed directly from acetyl-coenzyme A (CoA) through butyryl-CoA (hot channel). However, little is known about the relative contributions of the two butanol-forming pathways. Here we report that the direct butanol-forming pathway is a better channel to optimize for butanol production through metabolic flux and mass balance analyses. Butanol production through the hot channel was maximized by simultaneous disruption of the pta and buk genes, encoding phosphotransacetylase and butyrate kinase, while the adhE1(D485G) gene, encoding a mutated aldehyde/alcohol dehydrogenase, was overexpressed. The ratio of butanol produced through the hot channel to that produced through the cold channel increased from 2.0 in the wild type to 18.8 in the engineered BEKW(pPthlAAD(**)) strain. By reinforcing the direct butanol-forming flux in C. acetobutylicum, 18.9 g/liter of butanol was produced, with a yield of 0.71 mol butanol/mol glucose by batch fermentation, levels which are 160% and 245% higher than those obtained with the wild type. By fed-batch culture of this engineered strain with in situ recovery, 585.3 g of butanol was produced from 1,861.9 g of glucose, with the yield of 0.76 mol butanol/mol glucose and productivity of 1.32 g/liter/h. Studies of two butanol-forming routes and their effects on butanol production in C. acetobutylicum described here will serve as a basis for further metabolic engineering of clostridia aimed toward developing a superior butanol producer. IMPORTANCE Renewable biofuel is one of the answers to solving the energy crisis and climate change problems. Butanol produced naturally by clostridia has superior liquid fuel characteristics and thus has the potential to replace gasoline. Due to the lack of efficient genetic manipulation tools, however, strain improvement has been rather slow. Furthermore, complex metabolic characteristics of acidogenesis followed by solventogenesis in this strain have hampered development of engineered clostridia having highly efficient and selective butanol production capability. Here we report for the first time the results of systems metabolic engineering studies of two butanol-forming routes and their relative importances in butanol production. Based on these findings, a metabolically engineered Clostridium acetobutylicum strain capable of producing butanol to a high titer with high yield and selectivity could be developed by reinforcing the direct butanol-forming flux.

Project description:INTRODUCTION:Current metabolomics approaches to unravel impact of diet- or lifestyle induced phenotype variation and shifts predominantly deploy univariate or multivariate approaches, with a posteriori interpretation at pathway level. This however often provides only a fragmented view on the involved metabolic pathways. OBJECTIVES:To demonstrate the feasibility of using Goeman's global test (GGT) for assessment of variation and shifts in metabolic phenotype at the level of a priori defined pathways. METHODS:Two intervention studies with identified phenotype variations and shifts were examined. In a weight loss (WL) intervention study obese subjects received a mixed meal challenge before and after WL. In a polyphenol (PP) intervention study obese subjects received a high fat mixed meal challenge (61E% fat) before and after a PP intervention. Plasma samples were obtained at fasting and during the postprandial response. Besides WL- and PP-induced phenotype shifts, also correlation of plasma metabolome with phenotype descriptors was assessed at pathway level. The plasma metabolome covered organic acids, amino acids, biogenic amines, acylcarnitines and oxylipins. RESULTS:For the population of the WL study, GGT revealed that HOMA correlated with the fasting levels of the TCA cycle, BCAA catabolism, the lactate, arginine-proline and phenylalanine-tyrosine pathways. For the population of the PP study, HOMA correlated with fasting metabolite levels of TCA cycle, fatty acid oxidation and phenylalanine-tyrosine pathways. These correlations were more pronounced for metabolic pathways in the fasting state, than during the postprandial response. The effect of the WL and PP intervention on a priori defined metabolic pathways, and correlation of pathways with insulin sensitivity as described by HOMA was in line with previous studies. CONCLUSION:GGT confirmed earlier biological findings in a hypothesis led approach. A main advantage of GGT is that it provides a direct view on involvement of a priori defined pathways in phenotype shifts.

Dataset Information

Protein Acetylation and Butyrylation Regulate the Phenotype and Metabolic Shifts of the Endospore-forming Clostridium acetobutylicum.

Publications

Protein Acetylation and Butyrylation Regulate the Phenotype and Metabolic Shifts of the Endospore-forming <i>Clostridium acetobutylicum</i>.

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