title: Noncanonical Myo9b-RhoGAP Accelerates RhoA GTP Hydrolysis by a Dual-Arginine-Finger Mechanism.
abstract: The GTP hydrolysis activities of Rho GTPases are stimulated by GTPase-activating proteins (GAPs), which contain a RhoGAP domain equipped with a characteristic arginine finger and an auxiliary asparagine for catalysis. However, the auxiliary asparagine is missing in the RhoGAP domain of Myo9b (Myo9b-RhoGAP), a unique motorized RhoGAP that specifically targets RhoA for controlling cell motility. Here, we determined the structure of Myo9b-RhoGAP in complex with GDP-bound RhoA and magnesium fluoride. Unexpectedly, Myo9b-RhoGAP contains two arginine fingers at its catalytic site. The first arginine finger resembles the one within the canonical RhoGAP domains and inserts into the nucleotide-binding pocket of RhoA, whereas the second arginine finger anchors the Switch I loop of RhoA and interacts with the nucleotide, stabilizing the transition state of GTP hydrolysis and compensating for the lack of the asparagine. Mutating either of the two arginine fingers impaired the catalytic activity of Myo9b-RhoGAP and affected the Myo9b-mediated cell migration. Our data indicate that Myo9b-RhoGAP accelerates RhoA GTP hydrolysis by a previously unknown dual-arginine-finger mechanism, which may be shared by other noncanonical RhoGAP domains lacking the auxiliary asparagine.
authors.affiliation: National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Fengshuang
authors.initials: F
authors.lastname: Yi
authors.affiliation: State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China.
authors.forename: Ruirui
authors.initials: R
authors.lastname: Kong
authors.affiliation: National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China.
authors.forename: Jinqi
authors.initials: J
authors.lastname: Ren
authors.affiliation: State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhu
authors.affiliation: Key Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China.
authors.forename: Jizhong
authors.initials: J
authors.lastname: Lou
authors.affiliation: State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China; Department of Neurology, Center for Genetic Medicine, Lurie Cancer Center, Northwestern University Feinberg School of Medicine, 303 E Superior, Chicago, IL 60611 USA. Electronic address: jane-wu@northwestern.edu.
authors.forename: Jane Y
authors.initials: JY
authors.lastname: Wu
authors.affiliation: National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China. Electronic address: wfeng@ibp.ac.cn.
authors.forename: Wei
authors.initials: W
authors.lastname: Feng
mesh_terms: D001120:Arginine / Q000378:metabolism*; D002384:Catalysis; D020134:Catalytic Domain / Q000502:physiology; D005459:Fluorides / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D017616:Magnesium Compounds / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D020741:rho GTP-Binding Proteins / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
chemical_list: D020690:GTPase-Activating Proteins; D017616:Magnesium Compounds; C113469:myosin IXB; C085379:rho GTPase-activating protein; C031288:magnesium fluoride; D006160:Guanosine Triphosphate; D001120:Arginine; D009218:Myosins; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D005459:Fluorides
keywords: GTPase-activating proteins; Myo9b; Rho GTPases; RhoA; RhoGAP domain