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Isolation and Properties of Cytoplasmic α-Glycerol 3-Phosphate Dehydrogenase from the Pectoral Muscle of the Fruit Bat, Eidolon helvum.

Agboola, Femi Kayode and Thomson, Alan and Afolayan, Adeyinka (2003) Isolation and Properties of Cytoplasmic α-Glycerol 3-Phosphate Dehydrogenase from the Pectoral Muscle of the Fruit Bat, Eidolon helvum. Journal of Biochemistry and Molecular Biology, 36 (2). pp. 159-166.

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Abstract

Cytoplasmic α-glycerol-3-phosphate dehydrogenase from fruit-bat-breast muscle was purified by ion-exchange and affinity chromatography. The specific activity of the purified enzyme was approximately 120 units/mg of protein. The apparent molecular weight of the native enzyme, as determined by gel filtration on Sephadex G-100 was 59,500 ± 650 daltons; its subunit size was estimated to be 35,700 ± 140 by SDS-polyacrylamide gel electrophoresis. The true Michaelis-Menten constants for all substrates at pH 7.5 were 3.9 ± 0.7mM, 0.65 ± 0.05mM, 0.26 ± 0.06 mM, and 0.005 ± 0.0004 mM for L-glycerol-3-phosphate, NAD+, DHAP, and NADH, respectively. The true Michaelis-Menten constants at pH 10.0 were 2.30 ± 0.21 mM and 0.20 ± 0.01mM for L-glycerol-3-phosphate and NAD+, respectively. The turnover number, kcat, of the forward reaction was 1.9 ± 0.2 × 104s-1. The treatment of the enzyme with 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) under denaturing conditions indicated that there were a total of eight cysteine residues, while only two of these residues were reactive towards DTNB in the native enzyme. The overall results of the in vitro experiments suggest that α-glycerol-3-phosphate dehydrogenase of the fruit bat preferentially catalyses the reduction of dihydroxyacetone phosphate to glycerol-3-phosphate.

Item Type: Article
Journal or Publication Title: Journal of Biochemistry and Molecular Biology
Uncontrolled Keywords: Animal/bat flight ; Fruit bat ; Glycerol-3-phosphate dehydrogenase ; Pectoral/breast muscle
Subjects: Q Science > QH Natural history > QH301 Biology
Departments: Faculty of Health and Medicine > Biomedical & Life Sciences
ID Code: 9425
Deposited By: Dr Alan Thomson
Deposited On: 09 Jun 2008 14:07
Refereed?: Yes
Published?: Published
Last Modified: 26 Jul 2012 18:36
Identification Number:
URI: http://eprints.lancs.ac.uk/id/eprint/9425

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