Substitution of histidine for arginine-101 of dinitrogenase reductase disrupts electron transfer to dinitrogenase

Dinitrogenase reductase from Klebsiella pneumoniae strain UN1041 has a histidine residue substituted for arginine at position 101. The mutant dinitrogenase reductase was purified and characterized in order to determine the importance of arginine-101 in the interaction between dinitrogenase and dinit...

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Bibliographic Details
Published in:Biochemistry (Easton) 1989-02, Vol.28 (3), p.1206-1212
Main Authors: Lowery, Robert G, Chang, Christina L, Davis, Lawrence C, McKenna, Marie Claire, Stevens, Philip J, Ludden, Paul W
Format: Article
Language:English
Subjects:
Adenosine Triphosphate - metabolism
Adenosine Triphosphate - pharmacology
Arginine
Dinitrogenase Reductase
Electron Spin Resonance Spectroscopy
Electron Transport
Ferredoxins - genetics
Ferredoxins - metabolism
Histidine
Iron - metabolism
Kinetics
Klebsiella pneumoniae
Klebsiella pneumoniae - enzymology
Molybdoferredoxin - metabolism
Mutation
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Summary:Dinitrogenase reductase from Klebsiella pneumoniae strain UN1041 has a histidine residue substituted for arginine at position 101. The mutant dinitrogenase reductase was purified and characterized in order to determine the importance of arginine-101 in the interaction between dinitrogenase and dinitrogenase reductase during electron transfer. Purified dinitrogenase reductase from UN1041 is a dimer of 67 kDa, contains a functional 4Fe-4S cluster, undergoes a MgATP-dependent conformational change, and is competent for ATP hydrolysis uncoupled from substrate reduction in the presence of dinitrogenase. However, the mutant protein is unable to support the reduction of protons or acetylene by dinitrogenase. A 100-fold molar excess of Kp2 from UN1041 does not inhibit electron transfer from wild-type dinitrogenase reductase to dinitrogenase. It is concluded that the interaction of dinitrogenase reductase with dinitrogenase during reductant-independent ATP hydrolysis is different than the interaction between the two proteins during electron transfer; the substitution of histidine for arginine at position 101 disrupts only the latter interaction. The same conclusions are reached using wild-type dinitrogenase reductase which has been ADP-ribosylated at arginine-101.
ISSN:0006-2960
1520-4995
DOI:10.1021/bi00429a038