The Structure of an Unconventional HD-GYP Protein from Bdellovibrio Reveals the Roles of Conserved Residues in this Class of Cyclic-di-GMP Phosphodiesterases

School of Biosciences, University of Birmingham, Birmingham, United Kingdom.
mBio (Impact Factor: 6.79). 08/2011; 2(5). DOI: 10.1128/mBio.00163-11
Source: PubMed


Cyclic-di-GMP is a near-ubiquitous bacterial second messenger that is important in localized signal transmission during the control of various processes, including virulence and switching between planktonic and biofilm-based lifestyles. Cyclic-di-GMP is synthesized by GGDEF diguanylate cyclases and hydrolyzed by EAL or HD-GYP phosphodiesterases, with each functional domain often appended to distinct sensory modules. HD-GYP domain proteins have resisted structural analysis, but here we present the first structural representative of this family (1.28 Å), obtained using the unusual Bd1817 HD-GYP protein from the predatory bacterium Bdellovibrio bacteriovorus. Bd1817 lacks the active-site tyrosine present in most HD-GYP family members yet remains an excellent model of their features, sharing 48% sequence similarity with the archetype RpfG. The protein structure is highly modular and thus provides a basis for delineating domain boundaries in other stimulus-dependent homologues. Conserved residues in the HD-GYP family cluster around a binuclear metal center, which is observed complexed to a molecule of phosphate, providing information on the mode of hydroxide ion attack on substrate. The fold and active site of the HD-GYP domain are different from those of EAL proteins, and restricted access to the active-site cleft is indicative of a different mode of activity regulation. The region encompassing the GYP motif has a novel conformation and is surface exposed and available for complexation with binding partners, including GGDEF proteins.

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    • "Site-directed mutagenesis showed that alanine substitutions of the eight metal-ligating residues either abolished or significantly reduced the catalytic activity, indicating that all three metal ions contribute to catalysis, although a more structural role in co-ordination of the active site cannot be ruled out (Bellini et al., 2013). Consistent with the crystal structure of the HD-G_P protein Bd1817 reported earlier (Lovering et al., 2011), the conserved His 221 and Asp 222 that constitute the HD-GYP motif were shown to be directly involved in metal binding. Thus, the authors further clarify the role for several of the metal co-ordination residues, which have previously been shown to be essential for PDE activity and are highly conserved within the HD-GYP family (Ryan, 2013). "
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