Journal of Structural and Functional Genomics

, Volume 13, Issue 3, pp 177–183 | Cite as

Crystal structure of a catalytically active GG(D/E)EF diguanylate cyclase domain from Marinobacter aquaeolei with bound c-di-GMP product

  • Sergey M. Vorobiev
  • Helen Neely
  • Bomina Yu
  • Jayaraman Seetharaman
  • Rong Xiao
  • Thomas B. Acton
  • Gaetano T. Montelione
  • John F. Hunt
Article

Abstract

Recent studies of signal transduction in bacteria have revealed a unique second messenger, bis-(3′–5′)-cyclic dimeric GMP (c-di-GMP), which regulates transitions between motile states and sessile states, such as biofilms. C-di-GMP is synthesized from two GTP molecules by diguanylate cyclases (DGC). The catalytic activity of DGCs depends on a conserved GG(D/E)EF domain, usually part of a larger multi-domain protein organization. The domains other than the GG(D/E)EF domain often control DGC activation. This paper presents the 1.83 Å crystal structure of an isolated catalytically competent GG(D/E)EF domain from the A1U3W3_MARAV protein from Marinobacter aquaeolei. Co-crystallization with GTP resulted in enzymatic synthesis of c-di-GMP. Comparison with previously solved DGC structures shows a similar orientation of c-di-GMP bound to an allosteric regulatory site mediating feedback inhibition of the enzyme. Biosynthesis of c-di-GMP in the crystallization reaction establishes that the enzymatic activity of this DGC domain does not require interaction with regulatory domains.

Keywords

Diguanylate cyclase GG(D/E)EF domain Cyclic di-GMP X-ray crystal structure Structural genomics 

Abbreviations

ASU

Asymmetric unit

c-di-GMP

Bis-(3′–5′)-cyclic dimeric guanosine monophosphate

DGC

Diguanylate cyclase

GG(D/E)EF domain

Diguanylate cyclase domain

GTP

Guanosine triphosphate

I-site

Inhibitory site

IPTG

Isopropyl-1-thio-β-D-galactopyranoside

NESG

Northeast Structural Genomics Consortium

PDB

Protein Data Bank

PDE

Phosphodiesterase

PSI

NIH Protein Structure Initiative

RMSD

Root-mean-square-deviation

SAD

Single-wavelength anomalous diffraction

Supplementary material

10969_2012_9136_MOESM1_ESM.docx (60 kb)
Supplementary material 1 (DOCX 61 kb)

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Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Sergey M. Vorobiev
    • 1
  • Helen Neely
    • 1
  • Bomina Yu
    • 1
  • Jayaraman Seetharaman
    • 1
  • Rong Xiao
    • 2
    • 3
  • Thomas B. Acton
    • 2
    • 3
  • Gaetano T. Montelione
    • 2
    • 3
    • 4
  • John F. Hunt
    • 1
  1. 1.Department of Biological SciencesThe Northeast Structural Genomics Consortium Columbia UniversityNew YorkUSA
  2. 2.Department of Molecular Biology and BiochemistryCenter for Advanced Biotechnology and MedicinePiscatawayUSA
  3. 3.The Northeast Structural Genomics ConsortiumRutgers, The State University of New JerseyPiscatawayUSA
  4. 4.Robert Wood Johnson Medical SchoolUMDNJPiscatawayUSA

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