Isolation of n-ethylmaleimide-labelled phlorizin-sensitive d-glucose binding protein of brush border membrane from rat kidney cortex

Abstract

A glucose receptor with high affinity for phlorizin from isolated brush border of rat kidney was labelled specifically with N-[ 14C]ethylmaleimide and then extracted from the membranes. After the solubilization of the brush borders with sodium dodecyl sulphate the N-[ 14C]ethylmaleimide-labelled receptor protein was isolated and was found to have a molecular weight of approximately 30 000 as determined by sodium dodecyl sulphate-polyacrylamide gel disc electrophoresis. The receptor protein eluted from the sodium dodecyl sulphate-containing gels migrates as a single band on sodium dodecyl sulphate-free polyacrylamide gels. The receptor protein can also be released from the brush borders with low concentrations of sodium deoxycholate. Under these conditions the molecular weight of the N-[ 14C]ethylmaleimide-labelled receptor protein is approximately 60 000 in contrast to the protein component solubilized with sodium dodecyl sulphate. Since this detergent is known to dissociate the brush border membrane into its protein components, our results suggest that the phlorizin- sensitive glucose receptor protein has a molecular weight of about 30 000.