Summary: The haematological and neurological consequences of cobalamin deficiency define the essential role of this vitamin in key metabolic reactions. The identification of cubilin‐amnionless as the receptors for intestinal absorption of intrinsic factor‐bound cobalamin and the plasma membrane receptor for cellular uptake of transcobalamin bound cobalamin have provided a clearer understanding of the absorption and cellular uptake of this vitamin. As the genes involved in the intracellular processing of cobalamins and genetic defects of these pathways are identified, the metabolic disposition of cobalamins and the proteins involved are being recognized. The synthesis of methylcobalamin and 5′‐deoxyadenosylcobalamin, their utilization in conjunction with methionine synthase and methylmalonylCoA mutase, respectively, and the metabolic consequences of defects in these pathways could provide insights into the clinical presentation of cobalamin deficiency.

The transcobalamin (TC, TCII) receptor (TCblR) on the plasma membrane binds TC- cobalamin (Cbl) and internalizes the complex by endocytosis. This receptor was purified from human placental membranes by affinity chromatography. Tryptic digest of the protein extracted from a sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel and subjected to liquid chromatography/mass spectrometry identified 4 peptides that matched with a membrane protein in the data bank. TCblR belongs to the low-density lipoprotein receptor family, with 2 low-density lipoprotein receptor type A domains separated by a complement-like cysteine-rich region. The 282-amino acid sequence includes a signal peptide of 31 residues, extracellular domain of 198 residues, a transmembrane region of 21 residues, and a cytoplasmic domain of 32 residues. The binding of TC-Cbl does not require the cytoplasmic domain or its orientation in the plasma membrane because the recombinant extracellular domain binds TC-Cbl with high affinity and specificity. The protein is heavily glycosylated and accounts for the 58-kDa size by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The human gene first identified as 8D6A and more recently as CD 320 encoding TCblR is located at p13.2 on the short arm of chromosome 19, spans a length of 6.224 kb, and is composed of 5 exons and 4 introns.

In cerebral folate deficiency syndrome, the presence of autoantibodies against the folate receptor (FR) explains decreased folate transport to the central nervous system and the clinical response to folinic acid. Autoantibody crossreactivity with milk FR from different species prompted us to test the effect of a milk‐free diet. Intervention with a milkfree diet in 12 children (nine males, three females; mean age 6y [SD 4y 11mo], range 1‐19y), decreased autoantibody titer significantly from 2.08pmol of FR blocked per ml of serum (SD 2.1; range 0.24‐8.35) to 0.35pmol (SD 0.49; range 0‐1.32; p=0.012) over 3 to 13 months, whereas FR autoantibody titer increased significantly to 6.53 (SD 6.08; range 0.54‐14.07; p=0.013) in nine children who were reexposed to milk for 6 to 14 weeks. In 12 children on a normal diet (eight males, four females; mean age 5y 5mo [SD 4y 1mo], range 1y 6mo‐16y 4mo), the antibody titer increased significantly from 0.84pmol of FR blocked per ml (SD 0.39; range 0.24‐1.44) to 3.04pmol (SD 1.42; range 0.84‐6.01; p=0.001) over 10 to 24 months. Decreasing the autoantibody titer with a milk‐free diet in conjunction with folinic acid therapy may be advocated for these patients.

Elevated methylmalonic acid in five asymptomatic newborns whose fibroblasts showed decreased uptake of transcobalamin‐bound cobalamin (holo‐TC), suggested a defect in the cellular uptake of cobalamin. Analysis of TCblR/CD320, the gene for the receptor for cellular uptake of holo‐TC, identified a homozygous single codon deletion, c.262_264GAG (p.E88del), resulting in the loss of a glutamic acid residue in the low‐density lipoprotein receptor type A‐like domain. Inserting the codon by site‐directed mutagenesis fully restored TCblR function. Hum Mutat 31:1–6, 2010.