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Abstract
The bacterium Myxococcus xanthus uses a G protein cycle to dynamically regulate the leading/lagging pole polarity axis. The G protein MglA is regulated by its GTPase-activating protein (GAP) MglB, thus resembling Ras family proteins. Here, we show structurally and biochemically that MglA undergoes a dramatic, GDP-GTP-dependent conformational change involving a screw-type forward movement of the central β2-strand, never observed in any other G protein. This movement and complex formation with MglB repositions the conserved residues Arg53 and Gln82 into the active site. Residues required for catalysis are thus not provided by the GAP MglB, but by MglA itself. MglB is a Roadblock/LC7 protein and functions as a dimer to stimulate GTP hydrolysis in a 2:1 complex with MglA. In vivo analyses demonstrate that hydrolysis mutants abrogate Myxococcus' ability to regulate its polarity axis changing the reversal behaviour from stochastic to oscillatory and that both MglA GTPase activity and MglB GAP catalysis are essential for maintaining a proper polarity axis.
Highlights
IntroductionRan. By cycling between an inactive, GDP-bound and an active, GTP-bound conformation, they act as nucleotidedependent molecular switches regulating cellular functions including growth, polarity and differentiation (Vetter and Wittinghofer, 2001; Cox and Der, 2010)
Small G proteins of the Ras superfamily can be divided into five major subfamilies: Ras, Rho, Rab, Arf/Arl and Received: 24 May 2011; accepted: July 2011; published online: August 2011& 2011 European Molecular Biology OrganizationRan
Since the intrinsic rates of GTP hydrolysis and GDP/GTP exchange are very slow, their cycle is regulated by GTPase-activating proteins (GAPs) and guanine nucleotide exchange factors (GEFs), which increase the intrinsic rates by orders of magnitude (Bos et al, 2007)
Summary
Ran. By cycling between an inactive, GDP-bound and an active, GTP-bound conformation, they act as nucleotidedependent molecular switches regulating cellular functions including growth, polarity and differentiation (Vetter and Wittinghofer, 2001; Cox and Der, 2010). GAPs stimulate GTP hydrolysis by complementing and/or stabilizing the G protein catalytic site in the transition state. GTPase activation relies on correct positioning of a nucleophilic water molecule by a crucial glutamine (for Ras, Rho, Ran, Rab, Arf/Arl) or histidine (Sar, elongation factors) (Scheffzek and Ahmadian, 2005; Bos et al, 2007). Rho-, Ras-, Rab-, Arfand Arl-specific GAPs supply an arginine finger in trans, which stabilizes the catalytic glutamine of the G protein and neutralizes the negative charge developing in the transition state (Scheffzek and Ahmadian, 2005; Bos et al, 2007). Ga subunits of heterotrimeric G proteins contain the glutamine as well as an intrinsic arginine and stimulation by RGS proteins occurs by stabilization of the intrinsic catalytic machinery (Sprang, 1997; Sprang et al, 2007)
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