5-7 genes encoding ADH, DNA and amino acid sequence determination and analysis, polymorphism and allelic frequencies analysis, gene ADH2 possesses 2 allelic forms with Ile308 or Val308, expression of ADH2 alloenzymes in Escherichia coli
DNA and amino acid sequence determination and analysis, expression of His-tagged enzyme under the control of the strong constitutive Arxula adeninivorans-derived TEF1 promoter in auxotrophic Arxula adeninivorans strain G1214, Hansenula polymorpha strain RB11, and Saccharomyces cerevisiae strainSEY6210, using different expression modules for transformation, evaluation of effeciency, overview. Expression in Arxula adeninivorans is most effective
expression in Escherichia coli. The correct folding of the AdhC enzyme from hyperthermophilic Pyrococcus furiosus in mesophilic recombinant Escherichia coli is greatly influenced by the cultivation temperature. When grown at temperatures above the optimal growth temperature, Escherichia coli produces heat shock proteins to prevent protein aggregation. Heat shock proteins are known for their chaperonin activity, i.e., they help the protein folding and are responsible for an efficient protein quality control. When heated at 45°C for 2.5 h prior to induction, an increase of the activity is monitored compared to the standard cultivation at 37°C. The fast increase to 42°C yields more active enzyme than the slow increase to 42°C. This suggests that heat shock proteins either assist in the correct folding of the AdhC, or maybe even allow for resolubilization of (partially) denatured molecules. The cultivation at 45°C is not successful
expression in Pyrococcus furiosus from which the native aldehyde oxidoreductase (AOR) gene is deleted. A strain containing the Thermoanaerobacter ethanolicus AdhE in a synthetic operon with AdhA is constructed. The AdhA gene is amplified from Thermoanaerobacter sp. X514. The amino acid sequence of TxAdhA is identical to that of TeAdhA. Of the bacterial strains expressing the various heterologous AdhE genes, only those containing AdhE and AdhA from Thermoanaerobacter sp. produced ethanol above background. The Te-AdhEA strain containing both AdhE and AdhA produces the most ethanol (4.2 mM), followed by Te-AdhE (2.6 mM), AdhA (1.8 mM) and Tx-AdhE (1.5 mM). Ethanol and acetate are the only major carbon end-products from glucose under these conditions. For these four strains, the amount of ethanol produced per estimated glucose consumed is increased from the background level to 1.2, 1.0, 0.8 and 0.7 respectively
expression of rat ADH5 in an in vitro transcription/translation system, GFP-tagged ADH5 in COS cells, but no soluble ADH5 protein from heterologously expression in Escherichia coli cells with expression systems successfully used for other mammalian ADHs, including fused to glutathione-S-transferase
expression of Tx-AdhE in Pyrococcus furiosus from with the native aldehyde oxidoreductase (AOR) gene is deleted. Ethanol and acetate are the only major carbon end-products from glucose under these conditions. The amount of ethanol produced per estimated glucose consumed is increased from the background level 0.7 respectively
gene ADH3, subcloning in Escherichia coli strain DH5alpha, recombinant expression of His6-tagged enzyme in Echerichia coli strain BL21 (DE3), complementation of the HpADH3 mutant by an HpADH3 expression cassette fused to a strong constitutive promoter, the resulting strain produced a significantly increased amount of ethanol compared to the wild-type strain in a glucose medium, while in a xylose medium, the ethanol production is dramatically reduced in an HpADH3 overproduction strain compared to that in the wild-type strain, semi-quantitative RT-PCR analysis
gene encoding for ADH of the haloalkaliphilic archaeon Natronomonas pharaonis, which has a 1,068-bp open reading frame that encodes a protein of 355 amino acids, is cloned into the pET28b vector and is expressed in Escherichia coli
gene YIM1, cloned from Saccharomyces cerevisiae strain BY4742, DNA and amino acid sequence determination and analysis, recombinant expression in Saccharomyces cerevisiae strain INVSc1
1.1.1.1 alcohol dehydrogenase
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