Affiliation:
1. Department of Medicinal Chemistry
2. Department of Biology, University of Utah, Salt Lake City, Utah 84112
Abstract
ABSTRACT
Members of the saframycin/safracin/ecteinascidin family of peptide natural products are potent antitumor agents currently under clinical development. Saframycin MX1, from
Myxococcus xanthus
, is synthesized by a nonribosomal peptide synthetase, SafAB, and an
O
-methyltransferase, SafC, although other proteins are likely involved in the pathway. SafC was overexpressed in
Escherichia coli
, purified to homogeneity, and assayed for its ability to methylate a variety of substrates. SafC was able to catalyze the O-methylation of catechol derivatives but not phenols. Among the substrates tested, the best substrate for SafC was
l
-dihydroxyphenylalanine (
l
-dopa), which was methylated specifically in the 4′-O position (
k
cat
/
K
m
= 5.5 × 10
3
M
−1
s
−1
). SafC displayed less activity on other catechol derivatives, including catechol, dopamine, and caffeic acid. The more labile
l
-5′-methyldopa was an extremely poor substrate for SafC (
k
cat
/
K
m
= ∼2.8 × 10
−5
M
−1
s
−1
).
l
-Dopa thioester derivatives were also much less reactive than
l
-dopa. These results indicate that SafC-catalyzed 4′-O-methylation of
l
-dopa occurs prior to 5′-C-methylation, suggesting that 4′-O-methylation is likely the first committed step in the biosynthesis of saframycin MX1. SafC has biotechnological potential as a methyltransferase with unique regioselectivity.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
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