Subcellular Localization and Properties of Glyoxylate Cycle Enzymes in the Liver of Rats with Alloxan Diabetes

S. V. Volvenkin^*, V. N. Popov, and A. T. Eprintsev

School of Biology and Soil Sciences, Voronezh State University, Universitetskaya pl. 1, Voronezh, 394693 Russia; fax: (7-073) 2-789-755; E-mail: root@bc.vucnit.voronezh.su

^* To whom correspondence should be addressed.

Received November 20, 1998; Revision received March 23, 1999

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Key enzymes of the glyoxylate cycle (isocitrate lyase and malate synthetase) were found in the liver and kidney of rats suffering from alloxan diabetes. The activities of these enzymes in the liver were 0.080 and 0.0430 U/mg protein, respectively. Isocitrate lyase activity in the kidney was 0.030 U/mg protein, and that of the malate synthetase was 0.018 U/mg protein. Peroxisomal localization of the enzymes was shown. A novel malate dehydrogenase isoform was found in a liver of rats suffering from the alloxan diabetes. The isocitrate lyase was isolated by selective (NH₄)₂SO₄ precipitation and DEAE-Toyopearl chromatography. The resulting enzyme preparation had specific activity 6.1 U/mg protein, corresponding to 76.25-fold purification with 32.6% yield. The isocitrate lyase was found to follow the Michaelis--Menten kinetic scheme (K_m for isocitrate, 0.08 mM) and to be competitively inhibited by glucose 1-phosphate (K_i = 1.25 mM), succinate (K_i = 1.75 mM), and citrate (K_i = 1.0 mM); the pH optimum of the enzyme was 7.5 in Tris-HCl buffer.

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KEY WORDS: glyoxylate cycle, alloxan diabetes, isocitrate lyase, malate synthetase, malate dehydrogenase, subcellular localization, kinetic characteristics

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