Structural basis for the distinct roles of non-conserved Pro116 and conserved Tyr124 of BCH domain of yeast p50RhoGAP

• Published in: Cellular and molecular life sciences : CMLS 2024-12, Vol.81 (1), p.216, Article 216
• Main Authors: Shankar, Srihari, Chew, Ti Weng, Chichili, Vishnu Priyanka Reddy, Low, Boon Chuan, Sivaraman, J.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino acids; Binding; Biochemistry; Biomedical and Life Sciences; Biomedicine; Cell Biology; Conserved Sequence; Dimerization; Helices; Homology; Humans; Life Sciences; Lipids; Models, Molecular; Original; Original Article; Proline; Proline - chemistry; Proline - genetics; Proline - metabolism; Protein Binding; Protein Domains; Protein interaction; Proteins; Residues; rhoA GTP-Binding Protein - chemistry; rhoA GTP-Binding Protein - genetics; rhoA GTP-Binding Protein - metabolism; RhoA protein; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - chemistry; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Structure-function relationships; Tyrosine - chemistry; Tyrosine - genetics; Tyrosine - metabolism; Yeast; Yeasts
• ISSN: 1420-682X; 1420-9071; 1420-9071
• DOI: 10.1007/s00018-024-05238-8

p50RhoGAP is a key protein that interacts with and downregulates the small GTPase RhoA. p50RhoGAP is a multifunctional protein containing the BNIP-2 and Cdc42GAP Homology (BCH) domain that facilitates protein–protein interactions and lipid binding and the GAP domain that regulates active RhoA population. We recently solved the structure of the BCH domain from yeast p50RhoGAP ( Y BCH) and showed that it maintains the adjacent GAP domain in an auto-inhibited state through the β5 strand. Our previous WT Y BCH structure shows that a unique kink at position 116 thought to be made by a proline residue between alpha helices α6 and α7 is essential for the formation of intertwined dimer from asymmetric monomers. Here we sought to establish the role and impact of this Pro116. However, the kink persists in the structure of P116A mutant Y BCH domain, suggesting that the scaffold is not dictated by the proline residue at this position. We further identified Tyr124 (or Tyr188 in H BCH) as a conserved residue in the crucial β5 strand. Extending to the human ortholog, when substituted to acidic residues, Tyr188D or Tyr188E, we observed an increase in RhoA binding and self-dimerization, indicative of a loss of inhibition of the GAP domain by the BCH domain. These results point to distinct roles and impact of the non-conserved and conserved amino acid positions in regulating the structural and functional complexity of the BCH domain.