My research focuses on several enzymes expressed by the Gram positive bacteria Enterococci. These enzymes include the flavoproteins NADH peroxidase, NADH oxidase and alpha-glycero-phosphate oxidase. The enzymatic mechanisms of these three enzymes are being studied by a combination of traditional spectrophotometric measurements and the creation of site-directed mutants to dissect the enzymatic reactions.
The genetic regulation of glycerol metabolism in Enterococcus is also being studied. This involves the investigation of transcriptional control of the glp operon, which has been cloned. This involves measuring transcription (mRNA levels) and the use of reporter genes fused to the glp promoter. Another way in which glycerol metabolism is controlled is by phosphorylation of glycerol kinase, which causes activation of the kinase and hence controls the flux through the catabolic pathway. The gene for enterococcal glycerol kinase has been cloned and overexpressed in E. coli. This protein is now being used to study the control of enzyme activity within the cell. |
Recent Publications (selected):
Claiborne A, Yeh JI, Mallett TC, Luba J, Crane EJ 3rd, Charrier V, Parsonage D.: Protein-sulfenic acids: diverse roles for an unlikely player in enzyme catalysis and redox regulation. Biochemistry 38:15407-16 (1999).
Mallett TC, Parsonage D, Claiborne A.: Equilibrium analyses of the active-site asymmetry in enterococcal NADH oxidase: role of the cysteine-sulfenic acid redox center. Biochemistry 38:3000-11 (1999).
Parsonage D, Luba J, Mallett TC, Claiborne A.: The soluble alpha-glycerophosphate oxidase from Enterococcus casseliflavus. Sequence homology with the membrane-associated dehydrogenase and kinetic analysis of the recombinant enzyme. J. Biol. Chem. 273:23812-22 (1998).
Charrier V, Buckley E, Parsonage D, Galinier A, Darbon E, Jaquinod M, Forest E, Deutscher J, Claiborne A.: Cloning and sequencing of two enterococcal glpK genes and regulation of the encoded glycerol kinases by phosphoenolpyruvate-dependent, phosphotransferase system-catalyzed phosphorylation of a single histidyl residue. J. Biol. Chem. 272(22):14166-74 (1997).
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