Nitrate Reductase-Deficient Mutants in Barley

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Abstract
Nitrate reductase-deficient barley (H. vulgare L.) mutants were assayed for the presence of a functional Mo cofactor determined from the activity of the molybdoenzyme, xanthine dehydrogenase and for nitrate reductase-associated activities. Rocket immunoelectrophoresis was used to detect nitrate reductase cross-reacting material in the mutants. The cross-reacting material levels of the mutants ranged from 8 to 136% of the wild type and were correlated with their nitrate reductase-associated activities, except for nar 1c, which lacked all associated nitrate reductase activities but had 38% of the wild-type cross-reacting materia. The cross-reacting material of 2 nar 1 mutants, as well as nar 2a, Xno 18, Xno 19 and Xno 29, exhibited rocket immunoprecipitates that were similar to the wild-type enzyme indicating structural homology between the mutant and wild-type nitrate reductase proteins. The cross-reacting materials of the 7 remaining nar 1 alleles formed rockets only in the presence of purified wild-type nitrate reductase, suggesting structural modifications of the mutant cross-reacting materials. All nar 1 alleles and Xno 29 had xanthine dehydrogenase activity indicating the presence of functional Mol cofactors. Apparently, nar 1 is the structural gene for nitrate reductase. Mutants nar 2a, Xno 18 and Xno 19 lacked xanthine dehydrogenase activity and are considered to be Mol cofactor deficient mutants. Cross-reacting material was not detected in uninduced wild-type or mutant extracts, suggesting that nitrate reductase is synthesized de novo in response to nitrate.