Effects of site-directed mutation on the function of the chloroplast ATP synthase subunit.

Abstract The previous work in our lab showed that the spinach chloroplast ATP synthase e mutant with 3 amino acid residues deleted from the N-terminus had much lower ability to inhibit ATP hydrolysis and block proton leakage in comparison to a mutant with 1 or 2 residues deleted from the N-terminus. The present study aimed at determining whether there is special importance in the structure and function of the N-terminal third residue of the chloroplast e subunit. The leucine residue at the N-terminal third site (Leu3) of the spinach chloroplast e subunit was replaced with Ile, Phe, Thr, Arg, Glu or Pro by site-directed mutagenesis, forming mutants eL3I, eL3F, eL3T, eL3R, eL3E and eL3P, respectively. These e variants all showed lower abilities to inhibit ATP hydrolysis and to block proton leakage, as compared to the wild type e subunit (eWT). The abilities of mutants eL3I and eL3F to restore the ATP synthesis activity of reconstituted membranes were higher than those of eWT, but the abilities of the other e variants were lower than that of eWT. These results indicate that the hydrophobic and neutral characteristics of Leu3 of the chloroplast e subunit are very important for its ability to inhibit ATP hydrolysis and block proton leakage, and for the ATP synthesis ability of ATP synthase. [ABSTRACT FROM AUTHOR]