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Bioenergetics pp 295-302 | Cite as

The Purification and Properties of Glycerol-3-Phosphate Dehydrogenase in the Mitochondrial Inner Membrane

  • Qi-shui Lin
  • Shan-ping Shi
  • Jian-hua Liu

Summary

Glycerol-3-phosphate dehydrogenase (E.C.I. 1.99.5) was solubilized from rabbit skeletal muscle mitochondria by Triton X-100 and purified through hydroxyapatite column chromatography, DEAE-Sepharose CL-6B column chromatography and sucrose density gradient ultracentrifugation. The preparation was electrophoretically pure. The apparent molecular weight of the enzyme polypeptide was 69,000. There were 1.7 mg Triton X-100 and 26 μg phospholipid per mg protein of the preparation. Each molecule contained one molecule of FAD. The absorption spectra were studied after the Triton X-100 had been replaced by octylglucoside. It was observed that the absorption spectra altered upon the addition of the substrate, indicating that the reduction of the enzyme induced conformational changes. If the sulfhydryl groups of the enzyme were modified by iodoacetamide, two negative absorption peaks appeared at 256 nm and 305 nm, concomitant with the loss of enzyme activity. Chemical modification of amino side chain and tryptophan residues was also studied.

Keywords

Tryptophan Residue Dihydroxyacetone Phosphate Amino Side Chain Differential Absorption Spectrum Cytosol NADH 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Abbreviations

FAD

flavin adenline dinucleotide

NAD+

nicotinamide adenime dinucleotide

Tris

[(tris hydroxymethyl] -aminomethane)

DCPIP

dichlorophenolindolphenol

EDTA

ethylenediaminetetraacetic acid

Pi

inorganic phosphate

SDS

sodium dodecyl sulfate

TNBS

2, 4, 6-trinitrobenzene sufonic acid

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Copyright information

© Plenum Press, New York 1990

Authors and Affiliations

  • Qi-shui Lin
    • 1
  • Shan-ping Shi
    • 1
  • Jian-hua Liu
    • 1
  1. 1.Shanghai Institute of BiochemistryAcademia SinicaShanghaiChina

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