Assembly of subunit d (Vma6p) and G (Vma10p) and the NMR solution structure of subunit G (G1–59) of the Saccharomyces cerevisiae V1VO ATPase

Understanding the structural traits of subunit G is essential, as it is needed for V1VO assembly and function. Here solution NMR of the recombinant N- (G1–59) and C-terminal segment (G61–114) of subunit G, has been performed in the absence and presence of subunit d of the yeast V-ATPase. The data show that G does bind to subunit d via its N-terminal part, G1–59 only. The residues of G1–59 involved in d binding are Gly7 to Lys34. The structure of G1–59 has been solved, revealing an α-helix between residues 10 and 56, whereby the first nine- and the last three residues of G1–59 are flexible. The surface charge distribution of G1–59 reveals an amphiphilic character at the N-terminus due to positive and negative charge distribution at one side and a hydrophobic surface on the opposite side of the structure. The C-terminus exhibits a strip of negative residues. The data imply that G1–59–d assembly is accomplished by hydrophobic interactions and salt-bridges of the polar residues. Based on the recently determined NMR structure of segment E18–38 of subunit E of yeast V-ATPase and the presently solved structure of G1–59, both proteins have been docked and binding epitopes have been analyzed.