title: Structural insight into the mechanism of c-di-GMP hydrolysis by EAL domain phosphodiesterases.
abstract: Cyclic diguanylate (or bis-(3'-5') cyclic dimeric guanosine monophosphate; c-di-GMP) is a ubiquitous second messenger that regulates diverse cellular functions, including motility, biofilm formation, cell cycle progression, and virulence in bacteria. In the cell, degradation of c-di-GMP is catalyzed by highly specific EAL domain phosphodiesterases whose catalytic mechanism is still unclear. Here, we purified 13 EAL domain proteins from various organisms and demonstrated that their catalytic activity is associated with the presence of 10 conserved EAL domain residues. The crystal structure of the TBD1265 EAL domain was determined in free state (1.8 Å) and in complex with c-di-GMP (2.35 A), and unveiled the role of conserved residues in substrate binding and catalysis. The structure revealed the presence of two metal ions directly coordinated by six conserved residues, two oxygens of c-di-GMP phosphate, and potential catalytic water molecule. Our results support a two-metal-ion catalytic mechanism of c-di-GMP hydrolysis by EAL domain phosphodiesterases.
authors.affiliation: Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Anatoli
authors.initials: A
authors.lastname: Tchigvintsev
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Xu
authors.forename: Alexander
authors.initials: A
authors.lastname: Singer
authors.forename: Changsoo
authors.initials: C
authors.lastname: Chang
authors.forename: Greg
authors.initials: G
authors.lastname: Brown
authors.forename: Michael
authors.initials: M
authors.lastname: Proudfoot
authors.forename: Hong
authors.initials: H
authors.lastname: Cui
authors.forename: Robert
authors.initials: R
authors.lastname: Flick
authors.forename: Wayne F
authors.initials: WF
authors.lastname: Anderson
authors.forename: Andrzej
authors.initials: A
authors.lastname: Joachimiak
authors.forename: Michael Y
authors.initials: MY
authors.lastname: Galperin
authors.forename: Alexei
authors.initials: A
authors.lastname: Savchenko
authors.forename: Alexander F
authors.initials: AF
authors.lastname: Yakunin
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002384:Catalysis; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D008958:Models, Molecular; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D015290:Second Messenger Systems; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; C062025:bis(3',5')-cyclic diguanylic acid; D010727:Phosphoric Diester Hydrolases; D006152:Cyclic GMP