Abstract
Subunit a of the Escherichia coli ATP synthase, a 30 kDa integral membrane protein, was puri¢ed to homogeneity by a novel procedure incorporating selective extraction into a monophasic mixture of chloroform, methanol and water, followed by Ni-NTA chromatography in the mixed solvent. Pure subunit a was reconstituted with subunits b and c and phospholipids
to form a functional proton-translocating unit. Nuclear magnetic resonance (NMR) spectra of the pure subunit a in the mixed solvent show good chemical shift dispersion and demonstrate the potential of the solvent mixture for NMR studies of the large membrane proteins that are currently intractable in aqueous detergent solutions.
2003 Federation of European Biochemical Societies.
Published
by Elsevier B.V. All rights reserved.
Key words: ATP synthase; Subunit a;
Membrane protein puri¢cation;
Chloroform^methanol^water solvent; Proton translocation;
TROSY
Subunit a of the Escherichia coli ATP synthase, a 30 kDa integral membrane protein, was puri¢ed to homogeneity by a novel procedure incorporating selective extraction into a monophasic mixture of chloroform, methanol and water, followed by Ni-NTA chromatography in the mixed solvent. Pure subunit a was reconstituted with subunits b and c and phospholipids
to form a functional proton-translocating unit. Nuclear magnetic resonance (NMR) spectra of the pure subunit a in the mixed solvent show good chemical shift dispersion and demonstrate the potential of the solvent mixture for NMR studies of the large membrane proteins that are currently intractable in aqueous detergent solutions.
2003 Federation of European Biochemical Societies.
Published
by Elsevier B.V. All rights reserved.
Key words: ATP synthase; Subunit a;
Membrane protein puri¢cation;
Chloroform^methanol^water solvent; Proton translocation;
TROSY
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