title: Lincomycin biosynthesis involves a tyrosine hydroxylating heme protein of an unusual enzyme family.
abstract: The gene lmbB2 of the lincomycin biosynthetic gene cluster of Streptomyces lincolnensis ATCC 25466 was shown to code for an unusual tyrosine hydroxylating enzyme involved in the biosynthetic pathway of this clinically important antibiotic. LmbB2 was expressed in Escherichia coli, purified near to homogeneity and shown to convert tyrosine to 3,4-dihydroxyphenylalanine (DOPA). In contrast to the well-known tyrosine hydroxylases (EC 1.14.16.2) and tyrosinases (EC 1.14.18.1), LmbB2 was identified as a heme protein. Mass spectrometry and Soret band-excited Raman spectroscopy of LmbB2 showed that LmbB2 contains heme b as prosthetic group. The CO-reduced differential absorption spectra of LmbB2 showed that the coordination of Fe was different from that of cytochrome P450 enzymes. LmbB2 exhibits sequence similarity to Orf13 of the anthramycin biosynthetic gene cluster, which has recently been classified as a heme peroxidase. Tyrosine hydroxylating activity of LmbB2 yielding DOPA in the presence of (6R)-5,6,7,8-tetrahydro-L-biopterin (BH4) was also observed. Reaction mechanism of this unique heme peroxidases family is discussed. Also, tyrosine hydroxylation was confirmed as the first step of the amino acid branch of the lincomycin biosynthesis.
authors: Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic ; Central-European Technology Institute, Brno, Czech Republic ; Crop Research Institute, Drnovska Prague, Czech Republic.
authors: Jitka
authors: J
authors: Novotna
authors: Jana
authors: J
authors: Olsovska
authors: Petr
authors: P
authors: Novak
authors: Peter
authors: P
authors: Mojzes
authors: Radka
authors: R
authors: Chaloupkova
authors: Zdenek
authors: Z
authors: Kamenik
authors: Jaroslav
authors: J
authors: Spizek
authors: Eva
authors: E
authors: Kutejova
authors: Marketa
authors: M
authors: Mareckova
authors: Pavel
authors: P
authors: Tichy
authors: Jiri
authors: J
authors: Damborsky
authors: Jiri
authors: J
authors: Janata
mesh_terms: D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004295:Dihydroxyphenylalanine / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D006418:Heme / Q000737:chemistry / Q000378:metabolism; D006420:Hemeproteins / Q000235:genetics / Q000378:metabolism*; D006900:Hydroxylation; D007501:Iron / Q000737:chemistry / Q000378:metabolism; D008034:Lincomycin / Q000096:biosynthesis*; D005810:Multigene Family; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics; D014443:Tyrosine / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D006420:Hemeproteins; D011994:Recombinant Proteins; D014443:Tyrosine; D006418:Heme; D004295:Dihydroxyphenylalanine; D008034:Lincomycin; D007501:Iron; D014446:Tyrosine 3-Monooxygenase