Affiliation:
1. Institute of Biological Sciences, University of Wales, Aberystwyth SY23 3DD, Wales, United Kingdom
Abstract
ABSTRACT
We have cloned, sequenced, and heterologously expressed a periplasmic cytochrome
c
from a lupanine-utilizing
Pseudomonas putida
strain. Aerobic batch cultivation of
Escherichia coli
TB1 harboring the cytochrome
c
gene placed downstream of the
lac
promoter in pUC9 vector resulted in significant production of the holo-cytochrome
c
in the periplasm (∼4 mg of hemoprotein/liter of culture). The recombinant cytochrome
c
was purified to homogeneity and was found to be functional in accepting electrons from lupanine hydroxylase while catalyzing hydroxylation of lupanine. Comparison of the N-terminal amino acid sequence of the isolated cytochrome
c
with that deduced from the DNA sequence indicated that the signal sequence was processed at the bond position predicted by the SigPep program. The molecular size of the cytochrome
c
determined by electrospray mass spectrometry (9,595) was in precise agreement with that predicted from the nucleotide sequence.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
Cited by
4 articles.
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