Cloning, analysis, and overexpression of the gene encoding isobutylamine N-hydroxylase from the valanimycin producer, Streptomyces viridifaciens.
The flavoprotein isobutylamine N-hydroxylase (IBAH) catalyzes the oxidation of isobutylamine to isobutylhydroxylamine, a key step in the biosynthesis of the azoxy antibiotic valanimycin. By using oligonucleotide primers designed from peptide sequence information derived from native IBAH, a fragment of the gene (vlmH) encoding IBAH was amplified by PCR from a genomic library of the valanimycin-producing organism, Streptomyces viridifaciens MG456-hF10. The gene fragment was then employed as a probe to clone the entire vlmH gene from an S. viridifaciens genomic library. Overexpression of the vlmH gene in Escherichia coli gave a soluble protein that was purified to homogeneity. The purified protein exhibited the catalytic activity expected for IBAH. The deduced amino acid sequence of IBAH exhibited the greatest similarity to the Sox/DszC protein from Rhodococcus sp. strain IGT38, a flavoprotein involved in the oxidation of dibenzothiophene to the corresponding sulfone. Significant similarities were also found between the amino acid sequence of IBAH and those of the acyl coenzyme A dehydrogenases.
Parry
R J
RJ
Department of Chemistry, Rice University, Houston, Texas 77251, USA. parry@rice.edu
Li
W
W
Cooper
H N
HN
D000595:Amino Acid Sequence; D000818:Animals; D001391:Azo Compounds / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins; D001483:Base Sequence; D002082:Butylamines / Q000378:metabolism*; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D015870:Gene Expression; D006801:Humans; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics
D001391:Azo Compounds; D001426:Bacterial Proteins; D002082:Butylamines; D004269:DNA, Bacterial; C048267:valanimycin; C053521:isobutylamine; D006899:Mixed Function Oxygenases; C103668:VlmH protein, Streptomyces viridifaciens