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CYP79B1 from Sinapis alba converts tryptophan to indole-3-acetaldoxime
Ist Teil von
Archives of biochemistry and biophysics, 2003, Vol.409 (1), p.235-241
Ort / Verlag
United States: Elsevier Inc
Erscheinungsjahr
2003
Quelle
MEDLINE
Beschreibungen/Notizen
The cytochrome P450
CYP79B1 from
Sinapis alba has been heterologously expressed in
Escherichia coli and shown to catalyze the conversion of tryptophan to indole-3-acetaldoxime. Three expression constructs were made, one expressing the native protein and two expressing proteins with different N-terminal modifications. The native construct gave the highest yield as estimated by enzymatic activity per liter of culture. Spheroplasts of
E. coli expressing
CYP79B1 were reconstituted with the
Arabidopsis thaliana NADPH:cytochrome P450 reductase ATR1 heterologously expressed in
E. coli to obtain enzymatic activity. This indicates that the
E. coli electron-donating system, flavodoxin/flavodoxin reductase, does not support CYP79B1 activity. Recombinant CYP79B1 has a
K
m for tryptophan of
29±2
μM
and a
V
max of
36.5±0.7
nmol
h
−1(
ml
culture
)
−1
. The identity at the amino acid level of
CYP79B1 is, respectively, 93 and 84% to
CYP79B2 and
CYP79B3 from
A. thaliana, and 96% to CYP79B5 (Accession No.
AF453287) from
Brassica napus. The CYP79B subfamily of cytochromes P450 is likely to constitute a group of orthologous genes in the biosynthesis of indole glucosinolates.