The N-Terminal GTPase Domain of p190RhoGAP Proteins Is a PseudoGTPase

Structure
Amy L Stiegler, Titus J Boggon

Abstract

The pseudoGTPases are a rapidly growing and important group of pseudoenzymes. p190RhoGAP proteins are critical regulators of Rho signaling and contain two previously identified pseudoGTPase domains. Here we report that p190RhoGAP proteins contain a third pseudoGTPase domain, termed N-GTPase. We find that GTP constitutively purifies with the N-GTPase domain, and a 2.8-Å crystal structure of p190RhoGAP-A co-purified with GTP reveals an unusual GTP-Mg2+ binding pocket. Six inserts in N-GTPase indicate perturbed catalytic activity and inability to bind to canonical GTPase activating proteins, guanine nucleotide exchange factors, and effector proteins. Biochemical analysis shows that N-GTPase does not detectably hydrolyze GTP, and exchanges nucleotide only under harsh Mg2+ chelation. Furthermore, mutational analysis shows that GTP and Mg2+ binding stabilizes the domain. Therefore, our results support that N-GTPase is a nucleotide binding, non-hydrolyzing, pseudoGTPase domain that may act as a protein-protein interaction domain. Thus, unique among known proteins, p190RhoGAPs contain three pseudoGTPase domains.

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Citations

Sep 8, 2020·The FEBS Journal·Amy L Stiegler, Titus J Boggon
Dec 6, 2018·Biochemical Society Transactions·Amy L Stiegler, Titus J Boggon
Aug 15, 2019·Science Signaling·António J M RibeiroPatrick A Eyers

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