Linn T G, Greenleaf A L, Shorenstein R G, Losick R
Proc Natl Acad Sci U S A. 1973 Jun;70(6):1865-9. doi: 10.1073/pnas.70.6.1865.
The activity of the sigma subunit of the RNA polymerase of Bacillus subtilis decreases markedly during the first 2 hr of sporulation. Moreover, sigma activity remains deficient throughout the sporulation process and in dormant spores. The time course of changes in RNA polymerase during sporulation indicates that alterations in the core of RNA polymerase occur after the loss of sigma activity. Core RNA polymerase purified after the second and before the ninth hour of sporulation fails to respond to vegetative sigma subunit in vitro and contains variable amounts of a 110,000-dalton polypeptide in place of the beta' subunit. Core RNA polymerase purified from dormant spores has a subunit structure indistinguishable from vegetative core enzyme.RNA polymerase purified by antibody precipitation from an extract of a mixture of sporulating and excess vegetative cells separately labeled with two different radioisotopes contains beta' subunit and no 110,000-dalton polypeptide. However, RNA polymerase purified from sporulating bacteria in the absence of excess vegetative cells progressively loses the beta' subunit at each stage of purification even in the presence of the protease inhibitor, phenylmethyl sulfonyl fluoride. These findings suggest that the alteration of the beta' subunit is due to proteolysis in vitro.
枯草芽孢杆菌RNA聚合酶的σ亚基活性在芽孢形成的最初2小时内显著降低。此外,在整个芽孢形成过程以及休眠孢子中,σ活性一直不足。芽孢形成过程中RNA聚合酶变化的时间进程表明,RNA聚合酶核心的改变发生在σ活性丧失之后。在芽孢形成的第二小时之后和第九小时之前纯化的核心RNA聚合酶在体外对营养型σ亚基无反应,并且含有可变数量的110,000道尔顿的多肽,以取代β'亚基。从休眠孢子中纯化的核心RNA聚合酶的亚基结构与营养型核心酶无法区分。通过抗体沉淀从分别用两种不同放射性同位素标记的芽孢形成细胞和过量营养细胞混合物的提取物中纯化的RNA聚合酶含有β'亚基,而没有110,000道尔顿的多肽。然而,在没有过量营养细胞的情况下从芽孢形成细菌中纯化的RNA聚合酶,即使在存在蛋白酶抑制剂苯甲基磺酰氟的情况下,在纯化的每个阶段也会逐渐失去β'亚基。这些发现表明β'亚基的改变是由于体外蛋白水解作用。