AUTODIGESTION OF LEXA AND PHAGE-LAMDA REPRESSORS
AUTODIGESTION OF LEXA AND PHAGE-LAMDA REPRESSORS
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DOI：
10.1073/pnas.81.5.1375
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发表时间：
1984-01-01
期刊：
影响因子：
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通讯作者：
LITTLE, JW
中科院分区：
文献类型：
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作者：
LITTLE, JW
莱克萨阻遏物的蛋白水解裂解是大肠杆菌SOS调节系统去阻遏的早期步骤。在体内和体外的数据表明recA蛋白在这种特定的蛋白水解反应中的作用。在某些条件下，高度纯化的莱克萨蛋白的特异性体外切割可以在不存在recA蛋白的情况下发生。该自身消化反应与rec-A依赖性裂解反应裂解相同的丙氨酸-甘氨酸键。几条证据表明，这不是由于污染蛋白酶活性。自消化刺激碱性pH值。它发生在EDTA的存在下，但刺激的存在下，Ca 2+，Co 2+或Mg 2+的几倍。该反应似乎是一级反应，其速率在很宽的范围内与蛋白质浓度无关，这强烈表明它是分子内的。纯化的噬菌体阻遏物也在类似条件下分解，产生类似于recA蛋白作用产生的产物。噬菌体λ。如先前在recA催化的体外反应和体内反应中所观察到的，阻遏物以比莱克萨慢得多的速率分解。这两种类型的切割之间的相关性也延伸到与突变阻遏蛋白的反应;结合位点特异性，它表明自消化和recA依赖性切割遵循，至少部分，类似的反应途径。这些发现表明，特异性裂解的莱克萨蛋白可以催化的蛋白质本身，并建议recA蛋白发挥间接的刺激作用，也许作为一个变构效应，在recA依赖的反应，而不是直接作为一种蛋白酶。蛋白酶活性位点和recA-识别位点位于莱克萨蛋白的中心或COOH-末端部分，因为含有莱克萨蛋白的这些部分的胰蛋白酶片段可以参与这两个反应。
Proteolytic cleavage of lexA repressor is an early step in derepression of the SOS regulatory system of Escherichia coli. In vivo and in vitro data have indicated a role for recA protein in this specific proteolytic reaction. Under certain conditions, specific in vitro cleavage of highly-purified lexA protein can take place in the absence of recA protein. This autodigestion reaction cleaved the same alanine-glycine bond as did the rec-A-dependent cleavage reaction. Several lines of evidence argued that it was not due to a contaminating protease activity. Autodigestion was stimulated by alkaline pH. It occurred in the presence of EDTA but was stimulated several fold by the presence of Ca2+, Co2+ or Mg2+. The reaction appeared to be 1st-order, and its rate was independent of protein concentration over a wide range, strongly suggesting that it is intramolecular. Purified phage .lambda. repressor also broke down under similar conditions to yield products like those resulting from recA protein action. Phage .lambda. repressor broke down at a far slower rate than did lexA, as previously observed in the recA-catalyzed in vitro reaction and in vivo. This correlation between the 2 types of cleavage also extended to the reactions with mutant repressor proteins; taken together with the site specificity, it suggests that autodigestion and recA-dependent cleavage follow, at least in part, a similar reaction pathway. These findings indicate that specific cleavage of lexA protein can be catalyzed by the protein itself and suggest that recA protein plays an indirect stimulatory role, perhaps as an allosteric effector, in the recA-dependent reaction, rather than acting directly as a protease. The protease active site and the recA-recognition site lie in the central or COOH-terminal portion of the lexA protein, since a tryptic fragment containing these portions of lexA protein could take part in both reactions.