풀망둑(Acanthogobius hasta) 젖산탈수소효소의 특성

Abstract

The lactate dehydrogenase (EC 1.1.1.27, LDH) isozymes in tissues from Acanthogobius hasta were characterized by biochemical, immunochemical and kinetic methods. The activities of LDH in skeletal muscle and eye tissues were 65.30 and 53.25 units, but LDH activities in heart and liver tissues were very low. LDH/CS (EC 4.1.3.7, citrate synthase) in skeletal muscle was the highest as 22.29. Specific activities of LDH in brain, eye and skeletal muscle were 56.45, 38.04 and 11.0 units/mg, respectively. The LDH isozymes in tissues were separated by polyacrylamide gel electrophoresis after immunoprecipitation with antiserum against eye-specific and liver-specific . LDH isozymes were detected predominantly in skeletal muscle, brain and eye tissues, and isozyme was detected in heart. Anodal eye-specific and cathodal liver-specific were coexpressed in A. hasta. The eye-specific isozyme showed higher activity in eye tissue, but liver-specific isozyme showed lower activity in liver. As a result, one part of molecular structures in , and eye-specific and liver-specific were similar, but in were different with each other. Therefore the subunit A may be conservative in evolution, and the evolution of subunit B seems to be faster than that of subunit A. The LDH isozyme of skeletal muscle was purified in the fraction from elution with NAD+ containing buffer of affinity chromatography and eye-specific isozyme was eluted right after , so the structure of eye-specific isozyme is similar to . And LDH activity remained 35.22-43.47% as a result of the inhibition by pyruvate, the Michaelis-Menten constant values for pyruvate was 0.080-0.098 mM, and Vmax were 153.85 units, 35.09 units in skeletal muscle and eye, respectively. Also the isozyme was the thermo-stablest and was stabler than isozyme. The optimum pH of LDH was 6.5. The results mentioned above indicate that isozymes in tissues showed the properties between LDH isozyme as A. hasta was adapted to hypoxic conditions. Also LDH seems to function more effectively under anaerobic condition because LDH in skeletal muscle and eye tissues have high affinity for pyruvate.