Cozzani I, Barsacchi R, Dibenedetto G, Saracchi L, Falcone G
J Bacteriol. 1975 Sep;123(3):1115-23. doi: 10.1128/jb.123.3.1115-1123.1975.
L-Glutamate decarboxylase (GAD) activity of Clostridium perfringens (ATCC 8009) cells grown in various culture conditions was investigated. Remarkable variations of GAD level occur during the growth cycle in thioglycollate broth. These changes are affected by the pH of the culture medium. Addition of alkali to the culture media results in decrease of cell GAD activity, whereas increase of enzyme level occurs only in cells growing in unbuffered media. The results indicate that the mechanism regulating the GAD levels is sensitive to the changes of pH (or buffering substances) rather than to the steady pH values. Neither repression by glucose nor induction by L-glutamate was observed. Moreover, high concentrations of the free amino acid substrate in the culture media considerably decrease cell GAD activity, owing to the buffering effect of the amino acid. The molecular mechanism supporting the variations of GAD activity during the growth cycle of the cells were investigated and tentatively related to the structural and functional properties of the pure enzyme. It is shown that the drop of GAD activity during the lag phase is due to protein breakdown. Evidence is presented suggesting a control of protein degradation by its quaternary structure. Data are also reported supporting de novo synthesis of GAD during the late logarithmic phase of cell growth. Finally, the possible role of GAD as part of the pH regulation system of C. perfringens cells is discussed in relation both to physiologic conditions of the bacterial cell and to the molecular mechanisms regulating the GAD activity in vivo.
研究了在各种培养条件下生长的产气荚膜梭菌（ATCC 8009）细胞的L-谷氨酸脱羧酶（GAD）活性。在巯基乙酸盐肉汤的生长周期中，GAD水平发生显著变化。这些变化受培养基pH值的影响。向培养基中添加碱会导致细胞GAD活性降低，而酶水平的增加仅发生在未缓冲培养基中生长的细胞中。结果表明，调节GAD水平的机制对pH值（或缓冲物质）的变化敏感，而不是对稳定的pH值敏感。未观察到葡萄糖抑制或L-谷氨酸诱导。此外，由于氨基酸的缓冲作用，培养基中高浓度的游离氨基酸底物会显著降低细胞GAD活性。研究了支持细胞生长周期中GAD活性变化的分子机制，并初步将其与纯酶的结构和功能特性联系起来。结果表明，延迟期GAD活性的下降是由于蛋白质分解。有证据表明蛋白质降解受其四级结构的控制。还报告了支持细胞生长对数后期GAD从头合成的数据。最后，结合细菌细胞的生理条件和体内调节GAD活性的分子机制，讨论了GAD作为产气荚膜梭菌细胞pH调节系统一部分的可能作用。
