Glycerol is the main byproduct in ethanol production by Saccharomyces cerevisiae. In order to improve ethanol yield and the substrate conversion, a cassette about 4.5 kb for gene homologous recombination, gpd2Δ::PGK1(PT)-POS5-HyBR, was constructed and transformed into the haploid strain S. cerevisiae S1 (MATa) to replace the GPD2 gene by POS5 gene. The NADH kinase gene POS5 was successfully over expressed in the recombinant strain S. cerevisiae S3. Comparing with the parent strain, the recombinant strain S. cerevisiae S3 exhibited an 8% increase in ethanol production and a 33.64% decrease in glycerol production in the conical flask fermentation with an initiatory glucose concentration of 150 g/L. Overexpression of NADH kinase gene seems effective in reducing glycerol production and increasing ethanol yield.
Ethanol ; chemistry ; Fermentation ; Glycerol ; chemistry ; Industrial Microbiology ; Mitochondrial Proteins ; genetics ; metabolism ; Phosphotransferases (Alcohol Group Acceptor) ; genetics ; metabolism ; Saccharomyces cerevisiae ; genetics ; metabolism ; Saccharomyces cerevisiae Proteins ; genetics ; metabolism

Polythioesters newly emerged as a type of novel polymer and they have showed great potential for application in industries. In this study, genes of butyrate kinase (buk) and phosphotransbutyrylase (ptb) from Clostridium acetobutylicum, and poly (3-hydroxybutyrate) (PHB) synthase gene from Thiocapsa pfennigii were used for construction of a metabolic pathway to synthesize the polythioesters. When 3-mercaptopropionate and 3-hydroxybutyrate were fed, poly (3-mercaptopropoinate) [poly (3MP)] and poly(3-mercaptopropionate-co-3-hydroxybutyrate) [poly(3MP-co-3HB)] were synthesized by recombinant Escherichia coli JM109 (pBPP1) harboring the constructed metabolic pathway. Results indicated clearly that all these genes are necessary for the synthesis of poly(3MP) and poly(3MP-co-3HB).
3-Hydroxybutyric Acid ; chemistry ; 3-Mercaptopropionic Acid ; chemistry ; Acyltransferases ; genetics ; metabolism ; Escherichia coli ; genetics ; metabolism ; Gas Chromatography-Mass Spectrometry ; Models, Biological ; Molecular Weight ; Phosphate Acetyltransferase ; genetics ; metabolism ; Phosphotransferases (Carboxyl Group Acceptor) ; genetics ; metabolism ; Plasmids ; Polymers ; chemistry ; metabolism ; Spectrophotometry, Infrared

4-(Cytidine-5-diphospho) -2-C-methyl-D-erythritol kinase is a key enzyme in the biosynthesis pathway of terpenoids. According to the transcriptome database, the specific primers were designed and used in PCR. The bioinformatic analysis of the sequenced TwCMK gene was performed in several bioinformatics software. The Real-time fluorescence quantification polymerase chain reaction (RT-qPCR) were used to detect the expression levels of TwCMK from T. wilfordii after elicitor MeJA supplied. The results showed that the full length of TwCMK cDNA was 1 732 bp encoding 387 amino acids. The theoretical isoelectric point of the putative TwCMK protein was 5.79 and the molecular weight was about 42.85 kDa. MeJA stimulated the rising of TwCMK expression in suspension cell and signally impacted at 24 h. The research provides a basis for further study on the regulation of terpenoid secondary metabolism and biological synthesis.
Amino Acid Sequence ; Cloning, Molecular ; Computational Biology ; Gene Expression Regulation, Plant ; Models, Molecular ; Molecular Sequence Data ; Phosphotransferases (Alcohol Group Acceptor) ; chemistry ; genetics ; metabolism ; Phylogeny ; Plant Proteins ; chemistry ; genetics ; metabolism ; Sequence Alignment ; Tripterygium ; chemistry ; enzymology ; genetics

This review summarizes the recent developments on the regulation of human pyruvate dehydrogenase complex (PDC) by site-specific phosphorylation by four kinases. Mutagenic analysis of the three phosphorylation sites of human pyruvate dehydrogenase (E1) showed the site-independent mechanism of phosphorylation as well as site-independent dephosphorylation of the three phosphorylation sites and the importance of each phosphorylation site for the inactivation of E1. Both the negative charge and size of the group introduced at site 1 were involved in human E1 inactivation. Mechanism of inactivation of E1 was suggested to be site-specific. Phosphorylation of site 1 affected E1 interaction with the lipoyl domain of dihydrolipoamide acetyltransferase, whereas phosphorylation site 3 appeared to be closer to the thiamine pyrophosphate (TPP)-binding region affecting coenzyme interaction with human E1. Four isoenzymes of pyruvate dehydrogenase kinase (PDK) showed different specificity for the three phosphorylation sites of E1. All four PDKs phosphorylated sites 1 and 2 in PDC with different rates, and only PDK1 phosphorylated site 3. PDK2 was maximally stimulated by the reduction/acetylation of the lipoyl groups of E2. Presence of the multiple phosphorylation sites and isoenzymes of PDK is important for the tissue-specific regulation of PDC under different physiological conditions.
Acetylation ; Binding Sites ; Gene Expression Regulation, Enzymologic ; Human ; Isoenzymes/*metabolism ; Kinetics ; Mutagenesis, Site-Directed ; Mutation ; Oxidation-Reduction ; Phosphorylation ; Phosphotransferases/chemistry/genetics/*metabolism ; Protein Structure, Tertiary ; Pyruvate Dehydrogenase (Lipoamide)/metabolism ; Pyruvate Dehydrogenase Complex/chemistry/genetics/*metabolism ; Substrate Specificity ; Support, U.S. Gov't, P.H.S. ; Thiamine Pyrophosphate/metabolism