当前位置： SCI文献检索 > JOURNAL OF MOLECULAR BIOLOGY期刊下所有文献 > Transcription regulation in thermophilic bacteria: high resolution contact probing of Bacillus stearothermophilus and Thermotoga neapolitana arginine repressor-operator interactions.

Transcription regulation in thermophilic bacteria: high resolution contact probing of Bacillus stearothermophilus and Thermotoga neapolitana arginine repressor-operator interactions.

Abstract：

:Arginine-mediated regulation is remarkably well conserved in very divergent bacteria, and shows a number of unusual features that distinguish arginine regulation from other transcriptional control mechanisms. The arginine repressor subunit consists of a basic N-terminal DNA-binding domain, which belongs to the winged helix-turn-helix family, connected through a flexible linker to an acidic C-terminal domain responsible for binding of arginine and assembly of the high-affinity holohexamer, which binds an approximately 40 bp target. To gain further insight into the molecular details of arginine repressor-operator interactions we have established a high resolution contact map of the argC operator from Bacillus stearothermophilus, a moderate thermophilic Gram-positive bacterium, and the argR operator from Thermotoga neapolitana, a Gram-negative hyperthermophile, with the corresponding ArgR proteins. Enzymatic and chemical footprinting have been combined with missing contact, pre-modification, base substitution, and small ligand binding interference techniques to gather information on backbone and base-specific contacts with major and minor groove determinants of the operators. Wild-type and mutant argC operators have been compared for their interaction with the repressor, using both in vivo and in vitro approaches. Our results indicate that the operators of B. stearothermophilus and T. neapolitana consist of two ARG box-like sequences, 18 bp imperfect palindromes, separated by two and three base-pairs, respectively, and that the repressors from thermophilic origin establish base-specific contacts with two major groove segments and the intervening minor groove of each ARG box, all aligned on one face of the helix. In contrast, no specific contacts are established in the minor groove facing the repressor in the centre of the operator, nevertheless this region plays a crucial structural role in complex formation, as indicated by mutant studies. This picture is reminiscent of arginine repressor binding in Escherichia coli, and therefore reinforces the uniform view of arginine regulation, but also reveals a number of striking differences at particular positions of the boxes and in the length and base-pair composition of the spacer connecting two ARG boxes in the operator. These might be responsible, in part, for subtle but important functional and mechanistic differences in the way species-specific repressors interact with their cognate target sites. These variations are underlined by the different behaviour of the repressors from E. coli, B. stearothermophilus and T. neapolitana in their potential to bind heterologous operators, their requirement for arginine, and the resistance of complex formation to non-specific competitor DNA. Our findings are discussed in view of the crystal structure of the arginine repressor from B. stearothermophilus.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Song H,Wang H,Gigot D,Dimova D,Sakanyan V,Glansdorff N,Charlier D

doi

10.1006/jmbi.2001.5236

keywords:

subject

Has Abstract

pub_date

2002-01-18 00:00:00

pages

255-74

issue

eissn

0022-2836

issn

1089-8638

pii

S0022283601952362

journal_volume

315

pub_type

杂志文章

在线工具

Transcription regulation in thermophilic bacteria: high resolution contact probing of Bacillus stearothermophilus and Thermotoga neapolitana arginine repressor-operator interactions.