Premium
The internal rotenone‐insensitivae NADPH dehydrogenase contributes to malate oxidation by potato tuber and pea leaf mitochondria
Author(s) -
Agius Stephanie C.,
Bykova Natalia V.,
Igamberdiev Abir U.,
Møller Ian M.
Publication year - 1998
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1034/j.1399-3054.1998.1040306.x
Subject(s) - rotenone , nad+ kinase , submitochondrial particle , malate dehydrogenase , biochemistry , dehydrogenase , nadph dehydrogenase , pisum , malic enzyme , sativum , citrate synthase , nadh dehydrogenase , enzyme , biology , mitochondrion , chemistry , botany , protein subunit , gene , nitric oxide synthase
Inside‐out submitochondrial particles from both potato ( Solanum tuberosum L. cv. Bintje) tubers and pea ( Pisum sativum L. cv. Oregon) leaves possess three distinct dehydrogenase activities: Complex I catalyzes the rotenone‐sensitive oxidation of deamino‐NADH, ND in (NADPH) catalyzes the rotenone‐insensitive and Ca 2+ ‐dependent oxidation of NADPH and ND in (NADH) catalyzes the rotenone‐insensitive and Ca 2+ ‐independent oxidation of NADH. Diphenylene iodonium (DPI) inhibits complex I, ND in (NADPH) and ND in (NADH) activity with a K i of 3.7, 0.17 and 63 µ M , respectively, and the 400‐fold difference in K i between the two ND in made possible the use of DPI inhibition to estimate ND in (NADPH) contribution to malate oxidation by intact mitochondria. The oxidation of malate in the presence of rotenone by intact mitochondria from both species was inhibited by 5 µ M DPI. The maximum decrease in rate was 10–20 nmol O 2 mg −1 min −1 . The reduction level of NAD(P) was manipulated by measuring malate oxidation in state 3 at pH 7.2 and 6.8 and in the presence and absence of an oxaloacetate‐removing system. The inhibition by DPI was largest under conditions of high NAD(P) reduction. Control experiments showed that 125 µ M DPI had no effect on the activities of malate dehydrogenase (with NADH or NADPH) or malic enzyme (with NAD + or NADP + ) in a matrix extract from either species. Malate dehydrogenase was unable to use NADP + in the forward reaction. DPI at 125 µM did not have any effect on succinate oxidation by intact mitochondria of either species. We conclude that the inhibition caused by DPI in the presence of rotenone in plant mitochondria oxidizing malate is due to inhibition of ND in (NADPH) oxidizing NADPH. Thus, NADP turnover contributes to malate oxidation by plant mitochondria.