Presence of bound substrate in lactate dehydrogenase from carp liver.
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Date
2012-06
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Abstract
While attempting to purify UDP-galactose 4-epimerase from carp liver extract at pH 8.0, it was observed that the preparation even after dialysis could reduce NAD to NADH, interfering epimerase assay. The NAD reduction activity and the epimerase were co-eluted in a series of chromatographic steps. Mass spectrometric analysis of semi-purified fraction revealed that carp liver lactate dehydrogenase (LDH) contained bound lactate which was converted to pyruvate in the presence of NAD. The enzyme-bound lactate and the association with epimerase stabilized LDH from trypsin digestion and thermal inactivation at 45°C by factors of 2.7 and 4.2 respectively, as compared to substrate-free LDH. LDH and epimerase do not belong to any one pathway, but are the rate-limiting enzymes of two different pathways of carbohydrate metabolism. Typically, strongly associated enzymes work in combination, such as two enzymes of the same metabolic pathway. In that background, co-purification of LDH and epimerase as reloaded in this study was an unusual phenomenon.
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Keywords
Carp liver enzymes, Substrate bound lactate dehydrogenase, Epimerase-lactate dehydrogenase association, Protein-ligand interaction
Citation
Banerjee Nupur, Bhattacharyya Debasish. Presence of bound substrate in lactate dehydrogenase from carp liver. Indian Journal of Biochemistry & Biophysics. 2012 Jun; 49(3): 182-188.