Chiou S H, Lee H J, Huang S M, Chang G G
Laboratory of Crystallin Research, National Taiwan University, Taipei, Republic of China.
J Protein Chem. 1991 Apr;10(2):161-6. doi: 10.1007/BF01024779.
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.
为了阐明这些晶状体结构蛋白与其酶活性之间的关系，对从鸟类和爬行类动物中分离出的ε-晶体蛋白以及 authentic 乳酸脱氢酶（LDHs）进行了动力学比较。与先前鉴定的鸭ε-晶体蛋白相似，凯门鳄ε-晶体蛋白在聚丙烯酰胺凝胶上通过硝基蓝四唑染色和传统动力学分析检测时，似乎具有真正且稳定的 LDH 活性。还测定并比较了该ε-晶体蛋白催化反应中丙酮酸、L-乳酸、NAD⁺以及辅酶的三种结构类似物的动力学参数。尽管ε-晶体蛋白与鸡心脏 LDH 在结构上有相似之处，但通过天然同工酶电泳以及通过初速度和底物抑制研究进行的动力学分析，揭示了电荷和动力学性质上的差异。结果发现，对凯门鳄ε-晶体蛋白分析的动力学数据更符合类似于 authentic LDHs 和鸭ε-晶体蛋白的强制有序双底物双产物顺序机制。基于动力学分析以及与猪和鸡的 authentic 心脏型和肌肉型 LDHs 的比较，首次将凯门鳄ε-晶体蛋白确定为心脏型 LDH。
