N-TERMINAL PROTEIN-SEQUENCE ANALYSIS OF THE RABBIT ERYTHROCYTE LACTATE TRANSPORTER SUGGESTS IDENTITY WITH THE CLONED MONOCARBOXYLATE TRANSPORT PROTEIN MCT1

An improved purification for the rabbit erythrocyte lactate transporte r, using aminoethyl-Sepharose chromatography, is described. The proces s of purification of the 40-50 kDa transporter, labelled with 4,4'-dii sothiocyanostilbene-2,2'-disulphonate (DIDS), was followed by Western blotting with anti-DIDS antibodies [Poole, R. C. and Halestrap, A, P. (1992) Biochem. J. 283, 855-862]. Fractions highly-enriched in transpo rter were further purified by SDS/PAGE and the 40-50 kDa DIDS-labelled polypeptide was subjected to N-terminal protein sequencing. This anal ysis identified the first 16 amino acids of the protein. With the exce ption of one conservative substitution, this protein sequence is ident ical to the N-terminal protein sequence predicted from a cDNA isolated from Chinese hamster ovary cells that encode a monocarboxylate transp orter, MCT1 [Kim Garcia, C., Goldstein, J. L., Pathak, R.K., Anderson, R. G. W. and Brown, M. S. (1994) Cell 76, 865-873]. This observation, along with similarities in functional properties, leads us to conclud e that lactate transport in rabbit erythrocytes is mediated by the MCT 1 monocarboxylate transporter isoform.