Kinetic comparison of caiman epsilon-crystallin and authentic lactate dehydrogenases of vertebrates.
Kinetic comparison of epsilon-crystallins isolated from the avian and reptilian species and the authentic lactate dehydrogenases (LDHs) was undertaken in order to clarify the identities of these structural lens proteins in relation to their enzymatic activity. Caiman epsilon-crystallin similar to the previously characterized duck epsilon-crystallin appeared to possess a genuine and stable LDH activity as detected by nitro blue tetrazolium staining on polyacrylamide gels and conventional kinetic assays. Kinetic parameters for pyruvate, L-lactate, NAD+, and three structural analogues of the coenzyme in this epsilon-crystallin catalyzed reaction were also determined and compared. Despite the structural similarities between epsilon-crystallins and chicken heart LDH, differences in charge and kinetic properties have been revealed by native isozyme electrophoresis and kinetic analysis as examined by initial velocity and substrate inhibition studies. It is found that the kinetic data analyzed for caiman epsilon-crystallin were more fitted with a compulsory ordered Bi-Bi sequential mechanism similar to those for the authentic LDHs and duck epsilon-crystallin. Caiman epsilon-crystallin has for the first time been established as a heart-type LDH based on the kinetic analysis and comparison with the authentic heart- and muscle-type LDHs from pig and chicken.
Chiou
S H
SH
Laboratory of Crystallin Research, National Taiwan University, Taipei, Republic of China.
Lee
H J
HJ
Huang
S M
SM
Chang
G G
GG
D000489:Alligators and Crocodiles / Q000378:metabolism*; D000818:Animals; D002645:Chickens; D003459:Crystallins / Q000378:metabolism*; D004372:Ducks / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism*; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D007908:Lens, Crystalline / Q000201:enzymology*; D009132:Muscles / Q000201:enzymology; D009206:Myocardium / Q000201:enzymology; D009243:NAD / Q000378:metabolism; D009580:Nitroblue Tetrazolium; D011773:Pyruvates / Q000378:metabolism; D019289:Pyruvic Acid; D013194:Staining and Labeling; D013552:Swine
D003459:Crystallins; D007527:Isoenzymes; D007773:Lactates; D011773:Pyruvates; D009243:NAD; D009580:Nitroblue Tetrazolium; D019344:Lactic Acid; D019289:Pyruvic Acid; D007770:L-Lactate Dehydrogenase