Mapping the functional domains of TCblR/ CD320, the receptor for cellular uptake of transcobalamin-bound cobalamin.

The membrane receptor TCblR/CD320 binds transcobalamin (TC) saturated with vitamin B12 [cobalamin (Cbl)] and mediates cellular uptake of the vitamin. The specificity of TC for Cbl and of th receptor for TC-Cbl ensures efficient uptake of Cbl into cells. The high-affinity interaction of TCblR with TC-Cbl (K12= 10 nM-1 was investigated using deletions and mutations of amino acid sequences in TCblR. Only the extraceUular region (aa 32-229) is needed for TC-Cbl binding, but the N-glycosylation sites (N126, N195, and N213) are of no importance for this function. Deleting the cysteine-rich region (aa 95-141) that separates the two low-density lipoprotein receptor type A (LDLR-A) domains does not affect TC-Cbl binding (K12 = 19-24 nM-l). The two LDLR-A domains (aa 54-89 and 132-167) with the negatively charged acidic residues involved in Ca2+ binding are critical determinants of ligand binding. The cytoplasmic tail is apparently crucial for internalization of the ligand. Within this region, the RPL-GLL motif and the PDZ binding motifs (QERL/KESL appear to be involved in initiating and completing th process of ligand internalization. Mutations and deletions of these regions involved in binding and internalization of TC-Cbl are likely to produce the biochemical and clinical phenotype of Cbl deficiency. [ABSTRACT FROM AUTHOR]