ATP-dependent transcriptional activation by bacterial PspF AAA plus protein

被引:62
作者
Schumacher, J [1 ]
Zhang, XD [1 ]
Jones, S [1 ]
Bordes, P [1 ]
Buck, M [1 ]
机构
[1] Univ London Imperial Coll Sci Technol & Med, Dept Biol Sci, London SW7 2AZ, England
基金
英国生物技术与生命科学研究理事会;
关键词
PspF; enhancer-binding proteins; RNA polymerase; AAA proteins; transcription;
D O I
10.1016/j.jmb.2004.02.071
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Transcription activation by bacterial sigma(54)-depenclent enhancer-binding proteins (EBPs) requires their tri-nucleotide hydrolysis to restructure the sigma(54) RNA polymerase (RNAP). EBPs share sequence similarity with guanine nucleotide binding-proteins and ATPases associated with various cellular activities (AAA) proteins, especially in the mononucleotide binding P-loop fold. Using the phage shock protein F (PspF) EBP, we identify P-loop residues responsible for nucleotide binding and hydrolysis, consistent with their roles in other P-loop NTPases. We show the refined low-resolution structure of an EBP, PspF, revealing a hexameric ring organisation characteristic of AAA proteins. Functioning of EBPs involves ATP binding, higher oligomer formation and ATP hydrolysis coupled to the restructuring of the RNAP. This is thought to be a highly coordinated multi-step process, but the nucleotide-driven mechanism of oligomerisation and ATP hydrolysis is little understood. Our kinetic and structural data strongly suggest that three PspF dimers assemble to form a hexamer upon nucleotide binding. During the ATP hydrolysis cycle, both ATP and ADP are bound to oligomeric PspF, in line with a sequential hydrolysis cycle. We identify a putative R-finger, and show its involvement in ATP hydrolysis. Substitution of this arginine residue results in nucleotide-independent formation of hexameric rings, structurally linking the putative R-finger and, by inference, a specific nucleotide interaction to the control of PspF oligomerisation. (C) 2004 Elsevier Ltd. All rights reserved.
引用
收藏
页码:863 / 875
页数:13
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