Cysteine desulfurase activity indicates a role for NIFS in metallocluster biosynthesis.
Biological nitrogen fixation is catalyzed by nitrogenase, a complex metalloenzyme composed of two separately purifiable component proteins encoded by the structural genes nifH, nifD, and nifK. Deletion of the Azotobacter vinelandii nifS gene lowers the activities of both nitrogenase component proteins. Because both nitrogenase component proteins have metallocluster prosthetic groups that are composed of iron- and sulfur-containing cores, this result indicated that the nifS gene product could be involved in the mobilization of the iron or sulfur required for metallocluster formation. In the present work, it is shown that NIFS is a pyridoxal phosphate-containing homodimer that catalyzes the formation of L-alanine and elemental sulfur by using L-cysteine as substrate. NIFS activity is extremely sensitive to thiol-specific alkylating reagents, which indicates the participation of a cysteinyl thiolate at the active site. Based on these results we propose that an enzyme-bound cysteinyl persulfide that requires the release of the sulfur from the substrate L-cysteine for its formation ultimately provides the inorganic sulfide required for nitrogenase metallocluster formation. The recent discovery of nifS-like genes in non-nitrogen-fixing organisms also raises the possibility that the reaction catalyzed by NIFS represents a universal mechanism that involves pyridoxal phosphate chemistry, in the mobilization of the sulfur required for metallocluster formation.
Zheng
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Department of Anaerobic Microbiology, Virginia Polytechnic Institute and State University, Blacksburg 24061.
White
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Cash
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Jack
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D000595:Amino Acid Sequence; D016948:Azotobacter vinelandii / Q000201:enzymology* / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D013437:Carbon-Sulfur Lyases; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D003545:Cysteine / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D005033:Ethylmaleimide / Q000494:pharmacology; D005798:Genes, Bacterial; D007700:Kinetics; D008190:Lyases / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D005810:Multigene Family; D009586:Nitrogen Fixation / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008190:Lyases; D013437:Carbon-Sulfur Lyases; C080546:cysteine desulfurase; D003545:Cysteine; D005033:Ethylmaleimide