The proton-linked monocarboxylate transporters MCT1 – 4 play a crucial role in the shuttling of lactate between glycolytic and oxidative cell types in different tissues like muscle and brain. Using heterologous protein expression in Xenopus oocytes, we have recently reported that carbonic anhydrase isoform II (CAII), a ubiquitous enzyme catalysing the equilibration of carbon dioxide, protons and bicarbonate, enhances transport activity of MCT1. This interaction is independent of CAII catalytic activity, but seems to require physical interaction between transporter and enzyme (Becker & Deitmer 2008). We have now investigated potential binding between MCT and CA by co-expressing different isoforms and mutants of MCT and CA in Xenopus oocytes. Our results indicate that CAII enhances activity of MCT isoforms 1 and 4, but leaves MCT isoform 2 unaffected. Comparison of the protein sequence of MCT1, 2 and 4 revealed potential binding sites for CAII in the C-terminal of MCT1 and 4, but not in MCT2. For identification of the binding site, truncation mutants of MCT1 and single point mutations within the transporter's C-terminal were created. The mutation studies revealed that a cluster of three glutamate residues within the last 10 amino acids of MCT1 is crucial for the interaction between MCT1 and CAII. In contrast to the interaction between MCT1/4 with CAII, co-expression with CA isoform I did not enhance MCT activity. One prominent difference between CAI and CAII is a hisitidine-rich cluster within the N-terminal of CAII that is not found in CAI. As observed for CAI, co-expression of a CAII mutant lacking this cluster did not enhance activity of MCT1 and 4. Our results suggest that the modulation of MCT1/4 transport activity by CAII requires direct interaction between the proteins mediated by a glutamic acid cluster within the C-terminal of MCTs and a histidine cluster in the N-terminal of CAII.

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Becker, H.M. & Deitmer, J.W. 2008. J Biol Chem 283, 21655-67