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Review
. 2016 Aug;105(8):431-48.
doi: 10.1002/bip.22833.

Invited review: Small GTPases and their GAPs

Ashwini K Mishra  1 , David G Lambright  1
Affiliations
Review

Invited review: Small GTPases and their GAPs

Ashwini K Mishra et al. Biopolymers. 2016 Aug.

Abstract

Widespread utilization of small GTPases as major regulatory hubs in many different biological systems derives from a conserved conformational switch mechanism that facilitates cycling between GTP-bound active and GDP-bound inactive states under control of guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs), which accelerate slow intrinsic rates of activation by nucleotide exchange and deactivation by GTP hydrolysis, respectively. Here we review developments leading to current understanding of intrinsic and GAP catalyzed GTP hydrolytic reactions in small GTPases from structural, molecular and chemical mechanistic perspectives. Despite the apparent simplicity of the GTPase cycle, the structural bases underlying the hallmark hydrolytic reaction and catalytic acceleration by GAPs are considerably more diverse than originally anticipated. Even the most fundamental aspects of the reaction mechanism have been challenging to decipher. Through a combination of experimental and in silico approaches, the outlines of a consensus view have begun to emerge for the best studied paradigms. Nevertheless, recent observations indicate that there is still much to be learned. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 431-448, 2016.

Keywords: GAP; GTPase; hydrolysis; mechanism; structure.

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Figures

Figure 1
Figure 1. Small GTPase cycle and architecture
(A) GTPase cycle with key nucleotide exchange and GTP hydrolysis intermediates. Also indicated are metal fluoride complexes commonly used as ground and transition state mimetics in GAP complexes with GDP-bound small GTPases. (B) Structure of the prototypical small GTPase Ras in the active conformation bound to the non-hydrolyzeable GTP analog GppNHp (PDB ID 5P21). Characteristic G motifs, conformational switch regions and key functional residues are also indicated.
Figure 2
Figure 2. Reaction pathways and transition state intermediates for GTP hydrolysis
Transition state geometries corresponding to dissociative, associative, and concerted reaction pathways are depicted with approximate interatomic distances for the bond breakage (Pγ–Oβγ) and bond formation (Pγ–Owat) coordinates.
Figure 3
Figure 3. The arginine finger/cis-glutamine paradigm and variations
Hydrolytic sites in crystal structures of GTPases and GAP/GTPase complexes with ground or transition state mimetics. (A) Gtα-GDP-AlF4- (PDB ID 1TAD). Note Mg2+ is replaced by Ca2+ from the crystallization buffer. (B) RasGAP-Ras-GDP-AlF3 (PDB ID 1WQ1). (C) RasGAP-RasG12V-GDP-AlF3 with the G12V mutation modeled on the structure in B. (D) RhoGAP-Rho-GDP-MgF3- (PDB ID 1OW3). (E) RP2-Arl3-GDP-AlF4- (PDB ID 3BH7). (F) ASAP3-Arf6-GDP-AlF3 (PDB ID 3LVR).
Figure 4
Figure 4. Exceptions to the arginine finger/cis-glutamine paradigm
Hydrolytic sites in crystal structures of GAP/GTPase complexes with ground or transition state mimetics. (A) RanGAP-Ran-GDP-AlF3 (PDB ID 1K5G). (B) RapGAP-Rap-GDP-BeF3- (PDB ID 3BRW). (C) Gyp1-Rab33-GDP-AlF3 (PDB ID 2G77). (D) TBC1D20-Rab1-GDP-BeF3- (PDB ID 4HLQ). (E) EspG-Rab1-GDP-AlF3 (PDB ID 4FMC). (F) LepB-Rab1-GDP-AlF3 (PDB ID 4IRU).
Figure 5
Figure 5. Examples of GAP regulatory mechanisms
(A) Structure of autoinhibited β2-Chimaerin (PDB ID 1×A6) with Rho and phorbal ester derived from the structures of the RhoGAP complex with Rho-GDP-AlF4- (PDB ID 1TX4) and the PKCδ C1 domain bound to a phorbal ester (PDB ID 1PTR). (B) Structure of the ASAP3 PH domain with two molecules of dibutyl PIP2 simultaneously occupying the canonical and alternative phosphoinositide binding sites (PDB ID 5C79). The PH domain is shown in a hypothetical orientation relative to a simulated bilayer membrane. (C) Location of the switch II tyrosine AMPylated by Rab1 within the interface with LepB in the crystal structure of LepB-Rab1-GDP-AlF3 (PDB ID 4IRU). (D) Calcium binding site at the GAP-GTPase interface in the crystal structure of ASAP3-Arf6-GDP-AlF3 (PDB ID 3LVR).
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