Energy and calcium ion dependence of proteolysis during sporulation of Bacillus subtilis cells.
Energy and calcium ion dependence of proteolysis during sporulation of Bacillus subtilis cells.
复制标题
枯草芽孢杆菌细胞孢子形成过程中蛋白水解的能量和钙离子依赖性。
DOI：
10.1128/jb.172.8.4161-4170.1990
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发表时间：
1990
影响因子：
3.2
通讯作者：
Hageman,JH
中科院分区：
文献类型：
--
作者：
O'Hara,MB;Hageman,JH
细菌细胞在饥饿期间以升高的速率降解细胞内蛋白质，并且可以通过能量依赖性过程选择性地降解蛋白质。产孢细菌可以降解蛋白质的表观一级速率常数超过0.20 h-1。我们已经表明，优化的[14 C]亮氨酸标记和追逐程序，在化学成分确定的孢子形成培养基中，枯草芽孢杆菌168（trpC 2）的孢子形成细胞中的细胞内蛋白质降解显然是能量依赖性的。砷酸钠、叠氮化钠、羰基氰间氯苯腙和N，N '-二环己基碳二亚胺在孢子形成10小时内不引起明显溶解（小于或等于10%）的水平下，可抑制细胞内蛋白水解13 ~ 93%。指数生长的细胞获得砷酸盐抗性。与早期的报道相反，我们发现，氯霉素（100微克/毫升）强烈抑制蛋白水解（68%），甚至当添加到孢子形成过程6小时。限制培养基中的钙离子浓度（小于2 μ M）对营养生长的速率或程度没有影响，强烈抑制孢子形成（98%），并抑制60%或更多的蛋白水解率。能量代谢抑制剂在抑制蛋白水解的相同水平下，不影响细胞对Ca 2+的摄取速率或程度，这表明Ca 2+和蛋白水解的代谢能量需求是独立的。限制培养基中的Ca 2+浓度减少了三倍的细胞内的主要丝氨酸蛋白酶的比活性，在12小时的孢子形成后。最后，B的突变体的细胞。枯草杆菌携带的Ca 2（+）依赖性细胞内蛋白酶-1降解蛋白的插入失活基因在化学成分确定的孢子形成培养基中，以与野生型细胞无区别的速率培养8小时。
Bacterial cells degrade intracellular proteins at elevated rates during starvation and can selectively degrade proteins by energy-dependent processes. Sporulating bacteria can degrade protein with apparent first-order rate constants of over 0.20 h-1. We have shown, with an optimized [14C]leucine-labeling and chasing procedure, in a chemically defined sporulation medium, that intracellular protein degradation in sporulating cells of Bacillus subtilis 168 (trpC2) is apparently energy dependent. Sodium arsenate, sodium azide, carbonyl cyanide m-chlorophenylhydrozone, and N,N'-dicyclohexylcarbodiimide, at levels which did not induce appreciable lysis (less than or equal to 10%) over 10-h periods of sporulation, inhibited intracellular proteolysis by 13 to 93%. Exponentially growing cells acquired arsenate resistance. In contrast to earlier reports, we found that chloramphenicol (100 micrograms/ml) strongly inhibited proteolysis (68%) even when added 6 h into the sporulation process. Restricting the calcium ion concentration (less than 2 microM) in the medium had no effect on rates or extent of vegetative growth, strongly inhibited sporulation (98%), and inhibited rates of proteolysis by 60% or more. Inhibitors of energy metabolism, at the same levels which inhibited proteolysis, did not affect the rate or degree of uptake of Ca2+ by cells, which suggested that the Ca2+ and metabolic energy requirements of proteolysis were independent. Restricting the Ca2+ concentration in the medium reduced by threefold the specific activity in cells of the major intracellular serine proteinase after 12 h of sporulation. Finally, cells of a mutant of B. subtilis bearing an insertionally inactivated gene for the Ca2(+)-dependent intracellular proteinase-1 degraded protein in chemically defined sporulation medium at a rate indistinguishable from that of the wild-type cells for periods of 8 h.