Properties of submitochondrial particles from plant mitochondria: generation, surface characteristics and NAD(P)H oxidation

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Properties of submitochondrial particles from plant mitochondria : generation, surface characteristics and NAD(P)H oxidation. / Petit, Patrice X.; Gardeström, Per; Rasmusson, Allan G.; Møller, Ian M.

In: Plant Science, Vol. 78, No. 2, 1991, p. 177-183.

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T1 - Properties of submitochondrial particles from plant mitochondria

T2 - generation, surface characteristics and NAD(P)H oxidation

AU - Petit, Patrice X.

AU - Gardeström, Per

AU - Rasmusson, Allan G.

AU - Møller, Ian M.

PY - 1991

Y1 - 1991

N2 - Purified mitochondria isolated from potato (Solanum tuberosum L. cv. Bintje) tuber, Jerusalem artichoke (Helianthus tuberosu L.) tuber and rat liver were disrupted at different pH and different EDTA and MgCl2 concentrations either by French Press treatment or by sonication. The submitochondrial particles (SMP) were isolated by differential centrifugation and polarity estimated by the latency of cytochrome c oxidase (CCO) activity. The SMP were 5-9% inside-out depending on the conditions, and the disruption method was more important than the composition of the disruption medium in determining the polarity. At pH 6 and 7 and high-salt conditions sonication yielded SMP of the same polarity (82-91% inside-out) whereas French Press treatment in a low-salt buffer + EDTA gave more inside-out SMP at pH 6 than at pH 7. The inside out vesicles were able to build up a membrane potential in the presence of respiratory substrates (as tested with the anionic dye, oxonol VI) whereas no membrane potantial could be detected with the right-side-out vesicles (as tested with cationic dyes, and optical dye, safranine O, and a fluorescent dye, rhodamine 123) under similar conditions. Binding of Concanavalin A indicated that both the inner and outer surface of the inner membrane have exposed glycoproteins and/or glycolipids. Both right-side-out and inside-out SMP oxidized NADH, NADPH and succinate with good rates but there were clear differences in both donor and acceptor specificity between the outer and inner surface of the inner mitochondrial membrane. NADH oxidation by inside-out SMP was Ca2+-independent and rotenone-inhibited whereas NADPH oxidation by the inside-out SMP was Ca2+-dependent and relatively unaffected by rotenone.

AB - Purified mitochondria isolated from potato (Solanum tuberosum L. cv. Bintje) tuber, Jerusalem artichoke (Helianthus tuberosu L.) tuber and rat liver were disrupted at different pH and different EDTA and MgCl2 concentrations either by French Press treatment or by sonication. The submitochondrial particles (SMP) were isolated by differential centrifugation and polarity estimated by the latency of cytochrome c oxidase (CCO) activity. The SMP were 5-9% inside-out depending on the conditions, and the disruption method was more important than the composition of the disruption medium in determining the polarity. At pH 6 and 7 and high-salt conditions sonication yielded SMP of the same polarity (82-91% inside-out) whereas French Press treatment in a low-salt buffer + EDTA gave more inside-out SMP at pH 6 than at pH 7. The inside out vesicles were able to build up a membrane potential in the presence of respiratory substrates (as tested with the anionic dye, oxonol VI) whereas no membrane potantial could be detected with the right-side-out vesicles (as tested with cationic dyes, and optical dye, safranine O, and a fluorescent dye, rhodamine 123) under similar conditions. Binding of Concanavalin A indicated that both the inner and outer surface of the inner membrane have exposed glycoproteins and/or glycolipids. Both right-side-out and inside-out SMP oxidized NADH, NADPH and succinate with good rates but there were clear differences in both donor and acceptor specificity between the outer and inner surface of the inner mitochondrial membrane. NADH oxidation by inside-out SMP was Ca2+-independent and rotenone-inhibited whereas NADPH oxidation by the inside-out SMP was Ca2+-dependent and relatively unaffected by rotenone.

KW - calcium

KW - glycoprotein

KW - NAD(P)H oxidation

KW - plant mitochondria

KW - polarity

KW - submitochondrial particles

UR - http://www.scopus.com/inward/record.url?scp=3242697292&partnerID=8YFLogxK

U2 - 10.1016/0168-9452(91)90196-F

DO - 10.1016/0168-9452(91)90196-F

M3 - Article

VL - 78

SP - 177

EP - 183

JO - Plant Science

JF - Plant Science

SN - 0168-9452

IS - 2

ER -