Cloning of cDNAs encoding a rabbit renal brush border membrane protein immunologically related to band 3: Sequence similarity with microsomal dipeptidase

Distinct anion transport processes have been identified in the mammalian renal proximal tubule, but none of the responsible proteins or genes have been isolated. A 43kDa rabbit microvillus membrane protein that is immunologically related to the erythroid anion exchanger (band 3) was a candidate for a renal anion transporter. To examine the structural relationship with band 3, we cloned cDNAs encoding the 43kDa protein. The 43kDa band-3-1ike protein was purified, and a novel sequence of 24 amino acids was obtained from the N-terminus. Degenerate oligonucleotides were synthesized based on this sequence, and the polymerase chain reaction with single-sided specificity was used to amplify and clone a 1330bp cDNA from rabbit renal cortex. Additional overlapping 272bp and 1123bp cDNAs were obtained by synthesizing and screening a rabbit renal cortical cDNA library. The composite sequence was 1483bp, terminated with (A)₁₆, and was similar in size to the principal transcript expressed in rabbit renal cortex. The single long open reading frame was predicted to encode a protein composed of 410 amino acids with a molecular mass of 45193Da; 15 amino acids predicted to reside at the N-terminus were absent in the mature protein and may constitute a signal peptide. There was only limited sequence similarity with human erythroid band 3. Rather, the sequence was highly similar to microsomal dipeptidase, including the presence of a signal peptide and a consensus sequence for covalent linkage to glycosyl-phosphatidylinositol. In summary, the 43kDa protein from rabbit renal cortex that is recognized by a monospecific antibody to erythroid band 3 is most likely a microvillus membrane dipeptidase.