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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Cooperative Substrate Binding by a Diguanylate Cyclase

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Author(s):
Oliveira, Maycon C. [1] ; Teixeira, Raphael D. [1] ; Andrade, Maxuel O. [1] ; Pinheiro, Glaucia M. S. [2] ; Ramos, Carlos H. I. [2] ; Farah, Chuck S. [1]
Total Authors: 6
Affiliation:
[1] Univ Sao Paulo, Inst Chem, Dept Biochem, BR-05508070 Sao Paulo - Brazil
[2] Univ Estadual Campinas, Inst Chem, BR-13083970 Campinas, SP - Brazil
Total Affiliations: 2
Document type: Journal article
Source: Journal of Molecular Biology; v. 427, n. 2, p. 415-432, JAN 30 2015.
Web of Science Citations: 10
Abstract

XAC0610, from Xanthomonas citri subsp. citri, is a large multi-domain protein containing one GAF (cGMP-specific phosphodiesterases, adenylyl cyclases and FhlA) domain, four PAS (Per-Arnt-Sim) domains and one GGDEF domain. This protein has a demonstrable in vivo and in vitro diguanylate cyclase (DGC) activity that leads to the production of cyclic di-GMP (c-di-GMP), a ubiquitous bacterial signaling molecule. Analysis of a Xac Delta 0610 knockout strain revealed that XAC0610 plays a role in the regulation of Xac motility and resistance to H2O2. Site-directed mutagenesis of a conserved DGC lysine residue (Lys759 in XAC0610) resulted in a severe reduction in XAC0610 DGC activity. Furthermore, experimental and in silico analyses suggest that XAC0610 is not subject to allosteric product inhibition, a common regulatory mechanism for DGC activity control. Instead, steady-state kinetics of XAC0610 DGC activity revealed a positive cooperative effect of the GTP substrate with a dissociation constant for the binding of the first GTP molecule (K-1) approximately 5x greater than the dissociation constant for the binding of the second GTP molecule (K-2). We present a general kinetics scheme that should be used when analyzing DGC kinetics data and propose that cooperative GTP binding could be a common, though up to now overlooked, feature of these enzymes that may in some cases offer a physiologically relevant mechanism for regulation of DGC activity in vivo. (C) 2014 Elsevier Ltd. All rights reserved. (AU)

FAPESP's process: 11/07777-5 - Cyclic di-GMP signaling and the Type IV macromolecule secretion system in Xanthomonas citri
Grantee:Shaker Chuck Farah
Support Opportunities: Research Projects - Thematic Grants