Dissection of the functional and structural domains of phosphorelay histidine kinase a of Bacillus subtilis

Citation
L. Wang et al., Dissection of the functional and structural domains of phosphorelay histidine kinase a of Bacillus subtilis, J BACT, 183(9), 2001, pp. 2795-2802
Citations number
25
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Microbiology
Journal title
JOURNAL OF BACTERIOLOGY
ISSN journal
0021-9193 → ACNP
Volume
183
Issue
9
Year of publication
2001
Pages
2795 - 2802
Database
ISI
SICI code
0021-9193(200105)183:9<2795:DOTFAS>2.0.ZU;2-3
Abstract
The initiation of sporulation in Bacillus subtilis results primarily from p hosphoryl group input into the phosphorelay by histidine kinases, the major kinase being kinase A, Kinase A is active as a homodimer, the protomer of which consists of an approximately 100-amino-acid N-terminal putative signa l-sensing region and a 200-amino-acid C-terminal autokinase, On the basis o f sequence similarity, the N-terminal region may be subdivided into three P AS domains: A, B, and C, located from the N- to the C-terminal end. Proteol ysis experiments and two-hybrid analyses indicated that dimerization of the N-terminal region is accomplished through the PAS-B/PAS-C region of the mo lecule, whereas the most amino-proximal PAS-A domain is not dimerized, N-te rminal deletions generated with maltose binding fusion proteins showed that an intact PAS-A domain is very important for enzymatic activity. Amino aci d substitution mutations in PAS-A as well as PAS-C affected the in vivo act ivity of kinase A, suggesting that both PAS domains are required for signal sensing. The C-terminal autokinase, when produced without the N-terminal r egion, was a dimer, probably because of the dimerization required for forma tion of the four-helix-bundle phosphotransferase domain. The truncated auto kinase was virtually inactive in autophosphorylation with ATP, whereas phos phorylation of the histidine of the phosphotransfer domain by back reaction s from Spo0F similar toP appeared normal. The phosphorylated autokinase los t the ability to transfer its phosphoryl group to ADP, however. The N-termi nal region appears to be essential both for signal sensing and for maintain ing the correct conformation of the autokinase component domains.