McCormick et al. (J. Biol. Chem. 1993. 268:24683-24691) observed that mutations at ala-79 of the b subunit affect assembly of F1F0 ATP synthase. A polypeptide modeled on the soluble portion of the b subunit (bsol) has been used to investigate dimerization of the b subunit. Amino acid substitutions (ile,val,leu,pro,lys,gln,glu) were introduced into bsol at the position comparable toala-T9 of the b subunit using site-directed mutagenesis. ('hymotryptic digestion of the recombinant bsol polypeptides resulted in 15-kl)a fragments virtually identical to that observed for the wild type b subunit. The circular dichroism spectra of altered bsol proteins showed minima at 221 and 20Turn indicating predominately α-helical structures. Therefore, the gross structure of each mutated bsol appeared similar to the b subunit, ltowever, chemical crosslinking established that substitutions for ala-79 affected dimerization. The bsol ala-79→pro, gh, gln polypeptides were not cross-linked, while a reduced level of cross-linking was detected for' bsol ala-79→lys. Normal levels of cross-linking were detected for the bsol ala-79→val, leu, ile polypeptides. The data suggest that ala-79 is under structural constraints which are important tot dimeriza tiou of the hydrophilic domain of the b subunit. Comparison of these results with in vivo studies illustrate that loss of bsol dimerization call be correlated with biological phenotype.
|Original language||English (US)|
|State||Published - 1997|
ASJC Scopus subject areas
- Molecular Biology