Liu X, Matsumura P
Department of Microbiology and Immunology, University of Illinois at Chicago 60612-7344, USA.
Gene. 1995 Oct 16;164(1):81-4. doi: 10.1016/0378-1119(95)00480-t.
Based on the studies of the FliA protein in Bacillus subtilis (Bs) and Salmonella typhimurium (St), the Escherichia coli (Ec) fliA gene has been proposed to encode a flagellar-specific sigma factor, sigma 28. In this study, the complete nucleotide (nt) sequence of Ec fliA was determined. The fliA coding region consists of 717 nt starting with a GTG start codon and ending with a TAA stop codon. The gene product is predicted to be 239 amino acids (26,435 Da). Sequence comparison between Ec FliA and the sigma 28 of St revealed 93.7% identity. Gene fliA was amplified by the polymerase chain reaction, subcloned into expression vector pT7-7, and overexpressed. The overproduced 28-kDa FliA protein, recognized by the St anti-sigma 28 antibody, was purified to homogeneity. The purified protein was able to initiate transcription from the tar promoter in the presence of RNP core enzyme. We conclude that FliA functions as an alternative sigma factor sigma 28 which is specific for flagellar operons in Ec.
基于对枯草芽孢杆菌（Bs）和鼠伤寒沙门氏菌（St）中FliA蛋白的研究，有人提出大肠杆菌（Ec）的fliA基因编码一种鞭毛特异性的σ因子，即σ28。在本研究中，测定了Ec fliA的完整核苷酸（nt）序列。fliA编码区由717个核苷酸组成，起始于GTG起始密码子，终止于TAA终止密码子。预测该基因产物为239个氨基酸（26,435道尔顿）。Ec FliA与St的σ28之间的序列比较显示同一性为93.7%。通过聚合酶链反应扩增fliA基因，亚克隆到表达载体pT7-7中并进行过量表达。由St抗σ28抗体识别的过量产生的28 kDa FliA蛋白被纯化至同质。纯化后的蛋白能够在RNP核心酶存在的情况下从tar启动子起始转录。我们得出结论，FliA作为一种替代的σ因子σ28，对Ec中的鞭毛操纵子具有特异性。
