Götz F, Schleifer K H
Arch Microbiol. 1975 Nov 7;105(3):303-12. doi: 10.1007/BF00447150.
L-(+)-lactate dehydrogenase (LDH) from Staphylococcus epidermidis ATCC 14990 was purified by affinity chromatography. The purified enzyme was specifically activated by fructose-1,6-diphosphate (FDP). The concentration of FDP required for 50% maximal activity was about 0.15 mM. The enzyme activity was inhibited by adenosine diphosphate (ADP) and oxamate. The inhibition by ADP appeared to be competitive with respect to reduced nicotinamide adenine dinucleotide (NADH). The catalytic activity of the LDH for pyruvate reduction exhibited an optimum at pH 5.6. The enzyme is composed of four, probably identical, subunits. Sephadex gel filtration and sedimentation velocity at pH 5.6 Yielded molecular weights of about 130 000 and 126 000, respectively. The molecular weight at pH 6.5 and 7.0 was found to be only about 68 000. Polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate and sedimentation velocity at pH 2.0 or 8.5 revealed monomeric subunits with an approximate molecular weight of 36000. The thermostability of the heat labile enzyme was increased in the presence of FDP, NADH and pyruvate. The purified LDH exhibited an anomalous type of kinetic behavior. Plots of initial velocity vs. different concentrations of pyruvate, NADH or FDP led to saturation curves with intermediary plateau regions. As a consequence of these plateau regions the Hill coefficient alternated between lower and higher n-values. Some distinguishing properties of the S. epidermidis LDH and other LDHs activated by FDP are discussed.
通过亲和层析法纯化了来自表皮葡萄球菌ATCC 14990的L-(+)-乳酸脱氢酶（LDH）。纯化后的酶被1,6-二磷酸果糖（FDP）特异性激活。达到最大活性50%所需的FDP浓度约为0.15 mM。该酶的活性受到二磷酸腺苷（ADP）和草氨酸盐的抑制。ADP的抑制作用似乎对还原型烟酰胺腺嘌呤二核苷酸（NADH）具有竞争性。LDH催化丙酮酸还原的活性在pH 5.6时表现出最佳状态。该酶由四个可能相同的亚基组成。在pH 5.6条件下进行葡聚糖凝胶过滤和沉降速度分析，得到的分子量分别约为130000和126000。发现在pH 6.5和7.0时分子量仅约为68000。在十二烷基硫酸钠存在下进行聚丙烯酰胺凝胶电泳以及在pH 2.0或8.5条件下进行沉降速度分析，揭示出单体亚基的近似分子量为36000。在FDP、NADH和丙酮酸存在下，热不稳定酶的热稳定性增加。纯化后的LDH表现出一种异常的动力学行为。初始速度与不同浓度的丙酮酸、NADH或FDP的关系图导致具有中间平稳区域的饱和曲线。由于这些平稳区域，希尔系数在较低和较高的n值之间交替变化。讨论了表皮葡萄球菌LDH和其他被FDP激活的LDH的一些区别特性。
