Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase.

Journal article

Subunit a is a membrane-bound stator subunit of the ATP synthase and is essential for proton translocation. The N-terminus of subunit a in E. coli is localized to the periplasm, and contains a sequence motif that is conserved among some bacteria. Previous work has identified mutations in this region that impair enzyme activity. Here, an internal deletion was constructed in subunit a in which residues 6-20 were replaced by a single lysine residue, and this mutant was unable to grow on succinate minimal medium. Membrane vesicles prepared from this mutant lacked ATP synthesis and ATP-driven proton translocation, even though immunoblots showed a significant level of subunit a. Similar results were obtained after purification and reconstitution of the mutant ATP synthase into liposomes. The location of subunit a with respect to its neighboring subunits b and c was probed by introducing cysteine substitutions that were known to promote cross-linking: a_L207C + c_I55C, a_L121C + b_N4C, and a_T107C + b_V18C. The last pair was unable to form cross-links in the background of the deletion mutant. The results indicate that loss of the N-terminal region of subunit a does not generally disrupt its structure, but does alter interactions with subunit b.

Authors: Ishmukhametov, R; DeLeon-Rangel, J; Zhu, S; Vik, S

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Springer Verlag
• Journal: Journal of Bioenergetics and Biomembranes
• Volume: 49
• Issue: 2
• Pages: 171–181
• Publication date: 2017-01-11
• Acceptance date: 2017-01-04
• DOI: 10.1007/s10863-017-9694-z
• EISSN: 1573-6881
• ISSN: 0145-479X
• Language: English
• Keywords: ATPase; Proton translocation; Rotary motor; F1Fo; ATP synthase; Subunit a