In:
Plant Physiology, Oxford University Press (OUP), Vol. 120, No. 3 ( 1999-07-01), p. 841-848
Abstract:
The citrate carrier from maize(Zea mays) shoot mitochondria was solubilized with Triton X-100 and purified by sequential chromatography on hydroxyapatite and hydroxyapatite/celite in the presence of cardiolipin. SDS-gel electrophoresis of the purified fraction showed a single polypeptide band with an apparent molecular mass of 31 kD. When reconstituted into liposomes, the citrate carrier catalyzed a pyridoxal 5′-P-sensitive citrate/citrate exchange. It was purified 224-fold with a recovery of 50% and a protein yield of 0.22% with respect to the mitochondrial extract. In the reconstituted system the purified citrate carrier catalyzed a first-order reaction of citrate/citrate (0.065 min−1) or citrate/malate exchange (0.075 min−1). Among the various substrates and inhibitors tested, the reconstituted protein transported citrate,cis-aconitate, isocitrate, l-malate, succinate, malonate, glutarate, α-ketoglutarate, oxaloacetate, and α-ketoadipate and was inhibited by pyridoxal 5′-P, phenylisothiocyanate, mersalyl, andp-hydroxymercuribenzoate (but notN-ethylmaleimide), 1,2,3-benzentricarboxylate, benzylmalonate, and butylmalonate. The activation energy of the citrate/citrate exchange was 66.5 kJ/mol between 10°C and 35°C; the half-saturation constant (Km) for citrate was 0.65 ± 0.05 mm and the maximal rate (Vmax) of the citrate/citrate exchange was 13.0 ± 1.0 μmol min−1 mg−1 protein at 25°C.
Type of Medium:
Online Resource
ISSN:
1532-2548
,
0032-0889
DOI:
10.1104/pp.120.3.841
Language:
English
Publisher:
Oxford University Press (OUP)
Publication Date:
1999
detail.hit.zdb_id:
2004346-6
detail.hit.zdb_id:
208914-2
SSG:
12
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