Affiliation:
1. Department of Microbiology and Molecular Genetics, University of California, Los Angeles, California 90095-1489,1 and
2. Kluyver Department of Biotechnology, Delft University of Technology, 2628 BC Delft, The Netherlands2
Abstract
ABSTRACT
The nitrate reductase of the hyperthermophilic archaeon
Pyrobaculum aerophilum
was purified 137-fold from the cytoplasmic membrane. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, the enzyme complex consists of three subunits with apparent molecular weights of 130,000, 52,000, and 32,000. The enzyme contained molybdenum (0.8-mol/mol complex), iron (15.4-mol/mol complex) and cytochrome
b
(0.49-mol/mol complex) as cofactors. The
P. aerophilum
nitrate reductase distinguishes itself from nitrate reductases of mesophilic bacteria and archaea by its very high specific activity using reduced benzyl viologen as the electron donor (
V
max
with nitrate, 1,162 s
−1
(326 U/mg);
V
max
with chlorate, 1,348 s
−1
(378 U/mg) [assayed at 75°C]). The
K
m
values for nitrate and chlorate were 58 and 140 μM, respectively. Azide was a competitive inhibitor and cyanide was a noncompetitive inhibitor of the nitrate reductase activity. The temperature optimum for activity was >95°C. When incubated at 100°C, the purified nitrate reductase had a half-life of 1.5 h. This study constitutes the first description of a nitrate reductase from a hyperthermophilic archaeon.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
65 articles.
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