Control Of Alcohol Dehydrogenase Research Articles

Enzymatic synthesis of glutathione using engineered Saccharomyces cerevisiae

Two single gene cassettes, each containing one of the individual gene (γ-glutamylcysteine synthetase gene GSH1 or glutathione synthetase gene GSH2), were constructed under the control of alcohol dehydrogenase (ADH1) promoter and their respective native terminators. The recombinant plasmids constructed with Kan (r) or Hyg (r) as the selective markers and were transformed into Saccharomyces cerevisiae separately and jointly. Three engineered strains, GSH1-enhanced strain S.TS013/GSH1, GSH2-enhanced strain S.TS013/GSH2 and GSH1+GSH2 double-enhanced strain S.TS013/GSH1+GSH2, were constructed. Glutathione production using the recombinant strains to improve was then determined. By the cell dosage proportions of two engineered strains (S.TS013/GSH1, S.TS013/GSH2) and a two-stage reaction, GSH productivity increased by 84 and 59 % over that of the host strain and the S.TS013/GSH1+GSH2 strain, respectively.

Cytochrome P450 species specifically expressed in flower buds metabolize fatty acids

Cytochrome P450 (P450 or CYP) monooxygenases play an important role in the oxidative metabolism of a variety of lipophilic compounds including secondary metabolites as well as foreign chemicals in higher plants. It was reported that CYP78A1 was preferentially expressed in developing inflorescences of Zea mays. In addition, we isolated a cDNA clone of the novel cytochrome P450 species CYP703A1 from Petunia flower buds. A high level of the transcript of cyp703A1 was found at the early stage of flower development. In order to analyze the enzyme functions of these P450 species, CYP78A1 and CYP703A1 were expressed in the yeast Saccharomyces cerevisiae AH22 cells under the control of alcohol dehydrogenase I promoter and terminator. The recombinant yeast microsomes containing each of the CYP703A1 and CYP78A1 hemoproteins were found to metabolize lauric acid to give hydroxylated metabolites. We will discuss the relationship between the metabolism of fatty acids and flowering in both dicot and monocot plants.

Expression of rat liver microsomal cytochrome P-450 in yeast.

Recent advanced technologies of molecular cloning and protein chemistry revealed the entire primary structure of various cytochrome P-450 species so that the multiplicity of P-450 was evidently clarified. The domains responsible for heme binding and subcellular localization were posturated on the basis of the comparison of their primary structures.The rat microsomal P-450MC cDNA was efficiently expressed in the yeast Sacchromyces cerevisiae under the control of alcohol dehydrogenase I promoter and terminator. The P-450MC protein synthesized in yeast was localized in microsomes, contained heme in the molecule and interacted with an yeast electron-transport chain to exhibit monooxygenase activity.The yeast host-vector system combined with the site-directed mutagenesis and chimeric gene construction can be useful for overproduction and characterization of membrane-bound P-450s.

Genetical variation for enzyme activity in a population of Drosophila melanogaster. VI. Molecular variation in the control of alcohol dehydrogenase (ADH) activity.

Four characters, ADH activity at 25 degrees, immunologically determined ADH protein level, total protein and body weight were measured upon 72 hour old adult female and male Drosophila melanogaster from 16 highly inbred lines, derived from the laboratory population, "Texas" (established 1966). The highest levels of ADH activity and ADH protein level were observed in the 2 lined homozygous for the AdhF allele. Amongst the 14 AdhS/S lines variation for ADH protein level was associated with genetical variation for ADH activity (r = 0.6). The genetical association between ADH activity or ADH protein level and either body weight or total protein in the 16 inbred lines was not statistically significant. A study of ADH activity, ADH protein and total protein in 8 lines representing all homozygous combinations of chromosomes I, II and III and derived from two inbred AdhS/S lines, chosen for their respective high and low ADH activities, showed that ADH activity was considerably modified by a post-translational event controlled from chromosome III. Total protein was controlled by different chromosomal effects from those controlling ADH activity. Michaelis constants for crude fly extracts of the two AdhF/F and the above two AdhS/S lines showed clear differences in affinity for isopropanol.

Presumptive control mutation for alcohol dehydrogenase in Drosophila melanogaster

EXAMPLES of control mutations are rare in eukaryotes. Indeed, the only well-documented examples are in Aspergillus1,2 and a mutant affecting xanthine dehydrogenase (XDH) activity and mapping adjacent to, but separable from, rosy, the structural gene for XDH in Drosophila melanogaster3,4. Control of alcohol dehydrogenase (ADH), particularly in Drosophila has recently been studied. In examining ADH activity in selection lines and in samples from natural populations, a line was found to be Adhs and to have half the activity of a normal slow-migrating allele. To test the hypothesis that the low activity Adhs line contains a control mutation, we have attempted to separate the activity phenotype from the electrophoretic (structural gene) phenotype by recombination. We have succeeded in doing this, and in the process have obtained a very accurate position for Adh based on its recombination with closely-linked marker loci.

THE GENETIC CONTROL OF ALCOHOL DEHYDROGENASE IN MAIZE: GENE DUPLICATION AND REPRESSION

THE GENETIC CONTROL OF ALCOHOL DEHYDROGENASE IN MAIZE: GENE DUPLICATION AND REPRESSION