The thioesterase Bhp is involved in the formation of beta-hydroxytyrosine during balhimycin biosynthesis in Amycolatopsis balhimycina.
The putative hydrolase gene bhp from the balhimycin biosynthetic gene cluster has been cloned and overexpressed in Escherichia coli. The corresponding enzyme Bhp was purified to homogeneity by nickel-chelating chromatography and characterized. Although Bhp has sequence similarities to hydrolases with "haloperoxidase"/perhydrolase activity, it did not show any enzymatic activity with standard "haloperoxidase"/perhydrolase substrates (e.g., monochlorodimedone and phenol red), nonspecific esterase substrates (such as p-nitrophenyl acetate, p-nitrophenyl phosphate and S-thiophenyl acetate) or the model lactonase substrate dihydrocoumarin. However, Bhp could be shown to catalyse the hydrolysis of S-beta-hydroxytyrosyl-N-acetyl cysteamine thioester (beta-OH-Tyr-SNAC) with 15 times the efficiency of S-L-tyrosyl-N-acetyl cysteamine thioester (L-Tyr-SNAC). This is in agreement with the suggestion that Bhp is involved in balhimycin biosynthesis, during which it was supposed to catalyse the hydrolysis of beta-OH-Tyr-S-PCP (PCP=peptidyl carrier protein) to free beta-hydroxytyrosine (beta-OH-Tyr) and strongly suggests that Bhp is a thioesterase with high substrate specificity for PCP-bound beta-OH-Tyr and not a "haloperoxidase"/perhydrolase or nonspecific esterase.
Mulyani
Sri
S
Biochemistry, TU Dresden, 01062 Dresden, Germany.
Egel
Ellen
E
Kittel
Claudia
C
Turkanovic
Suada
S
Wohlleben
Wolfgang
W
Süssmuth
Roderich D
RD
van Pée
Karl-Heinz
KH
D000192:Actinomycetales / Q000201:enzymology* / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000737:chemistry; D055162:Biocatalysis; D004295:Dihydroxyphenylalanine / Q000378:metabolism*; D005810:Multigene Family; D013379:Substrate Specificity; D013869:Thiolester Hydrolases / Q000235:genetics / Q000378:metabolism*; D014640:Vancomycin / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000737:chemistry
D000900:Anti-Bacterial Agents; C087127:balhimycin; D004295:Dihydroxyphenylalanine; D014640:Vancomycin; D013869:Thiolester Hydrolases