Binding of sodium dodecyl suplhate to an integral membrane protein and to a water-soluble enzyme : Determination by molecular-sieve chromatography with flow scintillation detection

Abstract

We have determined the binding of sodium dodecyl (SDS) to the human red cell glucose transporter (polypeptide, M r 54 117) and to a water-soluble enzyme, N-5′-phosphoribosylanthranilate isomerase—indole-3-glycerol-phosphate synthase (PRAI—IGPS) from Escherichia coli ( M r 49 484). [ 35S]SDS was equilibrated with each protein on molecular-sieve chromatography at a series of SDS concentrations. The binding ratios of SDS to protein were determined by flow scintillationdetection and automated amino acid analyses. Unexpectedly the glucose transp an automated amino acid analyses. Unexpectedly the glucose transporter, which is a transmembrane protein, bound about the same amount of SDS per gram of protein as did the enzyme. At 1.6 m M SDS, slightly below the critical micelle concentration (CMC) (1.8 m M) in the eluent, the binding ratio was 1.6 g SDS/G protein for both the glucose transporter and PRAI—IGPS. At 2.0 m M SDS (above the CMC) the glucose transporter showed a binding ratio of 1.7 g SDS/g protein. The corresponding value for the enzyme was about 1.5 g/g. The SDS—glucose transporter complex seems to be more compact than the SDS—enzyme complex as judged by molecular-sieve chromatography and by SDS—polyacrylamide gel electrophoresis. Tecent neutron scattering results have shown a protein-decorated triple-micelle struct scattering results have shown a protein-decorated triple-micelle structure for the SDS—PRAI—IGPS complex. Hypothetically, the more compact SDS—glucose transporter complex may therefore consist of a dual-micelle structure. The molecular-sieve gel beads bound considerable amounts of SDS. The SDS binding to gel matrix and to the proteins increased with increasing SDS concentration up to at least 1.6–2.0 m M SDS. In the case of the water-soluble enzyme a shoulder was observed in the binding curve at 1 m M SDS, probably reflecting a change in the conformation of the complex.