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Plant Physiol, May 2000, Vol. 123, pp. 335-344
Ascorbate Biosynthesis in Mitochondria Is Linked to the Electron
Transport Chain between Complexes III and IV1
Carlos G.
Bartoli,
Gabriela M.
Pastori,* and
Christine H.
Foyer
Instituto de Fisiología Vegetal, Facultad de Ciencias
Agropecuarias, Universidad Nacional de La Plata, Casilla de Correos
327, (1900) La Plata, Argentina (C.G.B.); and Biochemistry and
Physiology Department, IACR-Rothamsted Experimental Station, Harpenden,
Herts AL5 2JQ, United Kingdom (G.M.P., C.H.F.)
Ascorbic acid is synthesized from galactono- -lactone (GL) in
plant tissues. An improved extraction procedure involving ammonium sulfate precipitation of membrane proteins from crude leaf homogenates yielded a simple, quick method for determining tissue activities of
galactono--lactone dehydrogenase (GLDH). Total foliar ascorbate and
GLDH activity decreased with leaf age. Subcellular fractionation experiments using marker enzymes demonstrated that 80% of the total
GLDH activity was located on the inner mitochondrial membrane, and 20%
in the microsomal fraction. Specific antibody raised against potato
(Solanum tuberosum L.) tuber GLDH recognized
a 56-kD polypeptide in extracts from the mitochondrial membranes but
failed to detect the equivalent polypeptide in microsomes. We
demonstrate that isolated intact mitochondria synthesize ascorbate in
the presence of GL. GL stimulated mitochondrial electron transport
rates. The respiration inhibitor antimycin A stimulated ascorbate
biosynthesis, while cyanide inhibited both respiration and ascorbate
production. GL-dependent oxygen uptake was observed in isolated intact
mitochondria. This evidence suggests that GLDH delivers electrons to
the mitochondrial electron transport chain between complexes III and IV.
1
C.B. acknowledges financial support from the
British Council, Consejo Nacional de Investigaciones
Científicas y Técnicas, and Fundacion Antorchas at
IACR-Rothamsted.
*
Corresponding author; e-mail gabriela.pastori{at}bbsrc.ac.uk;
fax
44-1582-763010.
© 2000 American Society of Plant Physiologists
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