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Transmembrane topology of Escherichia coli H + ‐ATPase (ATP synthase) subunit a
Author(s) -
Yamada Hiroshi,
Moriyama Yoshinori,
Maeda Masatomo,
Futai Masamitsu
Publication year - 1996
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(96)00621-7
Subject(s) - protein subunit , atp synthase gamma subunit , spheroplast , transmembrane protein , escherichia coli , v atpase , cytoplasm , atp synthase , biochemistry , membrane topology , transmembrane domain , polyclonal antibodies , microbiology and biotechnology , biology , atpase , peptide , topology (electrical circuits) , enzyme , membrane , atp hydrolysis , antibody , gene , receptor , mathematics , combinatorics , immunology
Escherichia coli H + ‐ATPase subunit a is a hydrophobic F 0 subunit. To investigate the topology of the subunit in the membrane, we prepared site‐specific polyclonal antibodies against amino‐terminal (Ser‐3 to Leu‐16), middle loop (Lys‐167 to Gln‐181), and carboxyl‐terminal (Thr‐259 to His‐271) peptide segments. Enzyme‐linked immunosorbent assay revealed that these antibodies specifically reacted with subunit a of inside‐out membrane vesicles, but not with that of right‐side‐out spheroplasts. Full reactivity appeared when spheroplasts were disrupted with Triton X‐100 (0.5%) or by sonication. These results suggest that at least parts of the three peptide segments of subunit a face the cytoplasm. Based on these observations, we propose a novel transmembrane topology of subunit a .