Biochemical Characterization of a Novel Indole Prenyltransferase from Streptomyces sp. SN-593
0301 basic medicine
Indoles
Magnetic Resonance Spectroscopy
Molecular Structure
Sequence Homology, Amino Acid
Molecular Sequence Data
Dimethylallyltranstransferase
Mass Spectrometry
Streptomyces
Substrate Specificity
Kinetics
03 medical and health sciences
Hemiterpenes
Organophosphorus Compounds
Bacterial Proteins
Amino Acid Sequence
Phylogeny
DOI:
10.1128/jb.01557-09
Publication Date:
2010-03-27T02:25:32Z
AUTHORS (8)
ABSTRACT
ABSTRACT
Genome sequencing of
Streptomyces
species has highlighted numerous potential genes of secondary metabolite biosynthesis. The mining of cryptic genes is important for exploring chemical diversity. Here we report the metabolite-guided genome mining and functional characterization of a cryptic gene by biochemical studies. Based on systematic purification of metabolites from
Streptomyces
sp. SN-593, we isolated a novel compound, 6-dimethylallylindole (DMAI)-3-carbaldehyde. Although many 6-DMAI compounds have been isolated from a variety of organisms, an enzyme catalyzing the transfer of a dimethylallyl group to the C-6 indole ring has not been reported so far. A homology search using known prenyltransferase sequences against the draft sequence of the
Streptomyces
sp. SN-593 genome revealed the
iptA
gene. The IptA protein showed 27% amino acid identity to cyanobacterial LtxC, which catalyzes the transfer of a geranyl group to (−)-indolactam V. A BLAST search against IptA revealed much-more-similar homologs at the amino acid level than LtxC, namely, SAML0654 (60%) from
Streptomyces ambofaciens
ATCC 23877 and SCO7467 (58%) from
S. coelicolor
A3(2). Phylogenetic analysis showed that IptA was distinct from bacterial aromatic prenyltransferases and fungal indole prenyltransferases. Detailed kinetic analyses of IptA showed the highest catalytic efficiency (6.13 min
−1
μM
−1
) for
l
-Trp in the presence of dimethylallyl pyrophosphate (DMAPP), suggesting that the enzyme is a 6-dimethylallyl-
l
-Trp synthase (6-DMATS). Substrate specificity analyses of IptA revealed promiscuity for indole derivatives, and its reaction products were identified as novel 6-DMAI compounds. Moreover, Δ
iptA
mutants abolished the production of 6-DMAI-3-carbaldehyde as well as 6-dimethylallyl-
l
-Trp, suggesting that the
iptA
gene is involved in the production of 6-DMAI-3-carbaldehyde.
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