Functional effects and cross-reactivity of antibody to purified subunit b (uncF protein) of Escherichia coli proton-ATPase |
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| Authors: | D S Perlin A E Senior |
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| Institution: | 1. Department of Neurology, The Affiliated Hospital, Xuzhou Medical University, Xuzhou, Jiangsu, 221002, China;2. Augusta University/University of Georgia Medical Partnership, Athens, GA, 30606, USA;3. Medical College of Georgia, Augusta University, Augusta, GA, 30912, USA;4. Brain Research Laboratory, Department of Emergency Medicine, Emory University School of Medicine, Atlanta, GA, 30032, USA;5. Key Laboratory of Anesthesiology of Jiangsu Province, Xuzhou, 221002, China;1. Nanotechnology and Catalysis Research Center (NanoCat), University Malaya, 50603 Kuala Lumpur, Malaysia;2. Department of Chemistry, Faculty of Science, University Malaya, 50603 Kuala Lumpur, Malaysia;3. Federal University Lafia Nasarawa State, PMB 146 Lafia, Nigeria;1. Servicio de Neumología, Complejo Asistencial Universitario de Salamanca, Salamanca, España;2. Servicio de Anatomía Patológica, Complejo Asistencial Universitario de Salamanca, Salamanca, España;3. Departamento de Medicina USAL, IBSAL, Salamanca, España |
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| Abstract: | Subunit b (uncF protein) of the proton-ATPase (F1F0) of Escherichia coli was purified from membranes of strain AN1460 (unc+). Antibody to purified subunit b was raised in rabbits. It reacted with F1-depleted membranes and blocked F1 binding. Bound antibody had no effect on proton transport through F0. F1-Depleted membranes competed with purified subunit b for antibody in an enzyme-linked immunosorbent assay. F1-Depleted membranes which had been pretreated with trypsin or preincubated with saturating amounts of soluble F1 competed poorly with purified subunit b for antibody. The antibody to subunit b was used to further evaluate the trypsin-cleavage data previously reported D. S. Perlin, D. N. Cox, and A. E. Senior (1983) J. Biol. Chem. 258, 9793-9800]. The results indicated that trypsin proteolysis of F1-depleted membranes resulted in the transient appearance of three fragments of subunit b (Mr = 16,400, 15,700, and 15,500) that remained tightly bound to the membrane. A water-soluble fragment (Mr 14,800), previously thought to be derived from subunit b, was not detected by the antibody. The antibody to subunit b did not cross-react with any subunit of mitochondrial, chloroplast, or other bacterial proton-ATPase, or with the proton-ATPase of clathrin-coated vesicles, plant microsomal membranes, or Neurospora crassa plasma membranes. |
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