Hansen Anne-Marie, Gu Yijun, Li Mi, Andrykovitch Michelle, Waugh David S, Jin Ding Jun, Ji Xinhua
Transcription Control Section, Gene Regulation and Chromosome Biology Laboratory, NCI-Frederick, National Institutes of Health, Frederick, Maryland 21702, USA.
J Biol Chem. 2005 Apr 29;280(17):17380-91. doi: 10.1074/jbc.M501444200. Epub 2005 Feb 25.
Stringent starvation protein A (SspA) of Escherichia coli is an RNA polymerase-associated transcriptional activator for the lytic development of phage P1 and is essential for stationary phase-induced acid tolerance of E. coli. We report the crystal structure of Yersinia pestis SspA, which is 83% identical to E. coli SspA in amino acid sequence and is functionally complementary in supporting the lytic growth of phage P1 and acid resistance of an E. coli sspA mutant. The structure reveals that SspA assumes the characteristic fold of glutathione S-transferase (GST). However, SspA lacks GST activity and does not bind glutathione. Three regions of SspA are flexible, the N and C termini and the alpha2-helix. The structure also reveals a conserved surface-exposed pocket composed of residues from a loop between helices alpha3 and alpha4. The functional roles of these structural features were investigated by assessing the ability of deletion and site-directed mutants to confer acid resistance of E. coli and to activate transcription from a phage P1 late promoter, thereby supporting the lytic growth of phage P1. The results indicate that the flexible regions are not critical for SspA function, whereas the surface pocket is important for both transcriptional activation of the phage P1 late promoter and acid resistance of E. coli. The size, shape, and property of the pocket suggest that it mediates protein-protein interactions. SspA orthologs from Y. pestis, Vibrio cholerae, and Pseudomonas aeruginosa are all functional in acid resistance of E. coli, whereas only Y. pestis SspA supports phage P1 growth.
大肠杆菌的严格饥饿蛋白A（SspA）是一种与RNA聚合酶相关的转录激活因子，参与噬菌体P1的裂解发育，对大肠杆菌在稳定期诱导的耐酸性至关重要。我们报道了鼠疫耶尔森菌SspA的晶体结构，其氨基酸序列与大肠杆菌SspA的同源性为83%，在支持噬菌体P1的裂解生长和大肠杆菌sspA突变体的耐酸性方面具有功能互补性。该结构显示，SspA具有谷胱甘肽S-转移酶（GST）的特征性折叠。然而，SspA缺乏GST活性，且不结合谷胱甘肽。SspA的三个区域具有柔性，即N端、C端和α2螺旋。该结构还揭示了一个由α3和α4螺旋之间的环上的残基组成的保守表面暴露口袋。通过评估缺失突变体和定点突变体赋予大肠杆菌耐酸性以及激活噬菌体P1晚期启动子转录的能力，从而支持噬菌体P1的裂解生长，研究了这些结构特征的功能作用。结果表明，柔性区域对SspA功能并不关键，而表面口袋对噬菌体P1晚期启动子的转录激活和大肠杆菌的耐酸性都很重要。口袋的大小、形状和性质表明它介导蛋白质-蛋白质相互作用。来自鼠疫耶尔森菌、霍乱弧菌和铜绿假单胞菌的SspA直系同源物在大肠杆菌的耐酸性方面均具有功能，而只有鼠疫耶尔森菌的SspA支持噬菌体P1生长。
