The δ subunil lias received very little attention in the study of ATP synthase preparations from eukaryotic cells. Studies reported here were designed both to provide a molecular description of the rat liver 8 subunit and to gain insight into the nature of its interaction withF1. The rat liver δ subunit was cloned from a λg11 library, sequenced, and overexpressed in E. coli and purified to homogeneity- The purified δ subunit (MW = 14,725 kDa) was to be highly structured and to exhibit about 25% sequence identity to the chloroplast and E. coli epsilon subunits, frequently regarded as homologs of higher eukaryotic δ subunits. In contrast to the chloroplast and £. coli epsilon subunits, which arc readily removed from their parent F1 moieties after treatment respectively with ethanol and lauryldimethy lamine oxide, the rat liver δ subunit remained tightly bound to F1 under these relatively mild conditions. For these reasons, experiments were carried out on rat liver F1 using antibodies, specific proteases, cross-linking agents, and selective treatment with SDS in order to gain insight into the nature of the interaction of the δ subunit with the intact enzyme complex. The data: a) supports the view that the rat liver F1δsubunit is in very close proximity to the γ subunit near the bottom of the F1 molecule but does not penetrate deeply into the central core; b) shows that within F1 the δ subunit's N-terminus is exposed while its C-tenninus is masked; c) indicates that access to the δ subunit is shielded in part by the α, β, and γ subunits and changes during the catalytic cycle of F1; and d) implicates the δ subunit as important for the structural stability of the F1 unit. These novel findings on a higher eukaiyotic F1 δ subunit will be summarized within the context of a model and discussed in relationship to earlier studies on the related epsilon subunits from both chloroplasts and E. coli.
|Original language||English (US)|
|State||Published - Dec 1 1998|
ASJC Scopus subject areas
- Molecular Biology