A Structural Network Analysis of Neuronal ArhGAP21/23 Interactors by Computational Modeling

[Image: see text] RhoGTPase-activating proteins (RhoGAPs) play multiple roles in neuronal development; however, details of their substrate recognition system remain elusive. ArhGAP21 and ArhGAP23 are RhoGAPs that contain N-terminal PDZ and pleckstrin homology domains. In the present study, the RhoGAP domain of these ArhGAPs was computationally modeled by template-based methods and the AlphaFold2 software program, and their intrinsic RhoGTPase recognition mechanism was analyzed from the domain structures using the protein docking programs HADDOCK and HDOCK. ArhGAP21 was predicted to preferentially catalyze Cdc42, RhoA, RhoB, RhoC, and RhoG and to downregulate RhoD and Tc10 activities. Regarding ArhGAP23, RhoA and Cdc42 were deduced to be its substrates, whereas RhoD downregulation was predicted to be less efficient. The PDZ domains of ArhGAP21/23 possess the FTLRXXXVY sequence, and similar globular folding consists of antiparalleled β-sheets and two α-helices that are conserved with PDZ domains of MAST-family proteins. A peptide docking analysis revealed the specific interaction of the ArhGAP23 PDZ domain with the PTEN C-terminus. The pleckstrin homology domain structure of ArhGAP23 was also predicted, and the functional selectivity for the interactors regulated by the folding and disordered domains in ArhGAP21 and ArhGAP23 was examined by an in silico analysis. An interaction analysis of these RhoGAPs revealed the existence of mammalian ArhGAP21/23-specific type I and type III Arf- and RhoGTPase-regulated signaling. Multiple recognition systems of RhoGTPase substrates and selective Arf-dependent localization of ArhGAP21/23 may form the basis of the functional core signaling necessary for synaptic homeostasis and axon/dendritic transport regulated by RhoGAP localization and activities.