Crystal structure of human intrinsic factor: Cobalamin complex at 2.6-Å resolution

Abstract

The structure of intrinsic factor (IF) in complex with cobalamin (Cbl) was determined at 2.6-Å resolution. The overall fold of the molecule is that of an ␣6/␣6 barrel. It is a two-domain protein, and the Cbl is bound at the interface of the domains in a base-on conformation. Surprisingly, two full-length molecules, each comprising an ␣-and a ␤-domain and one Cbl, and two truncated molecules with only an ␣-domain are present in the same asymmetric unit. The environment around Cbl is dominated by uncharged residues, and the sixth coordinate position of Co 2؉ is empty. A detailed comparison between the IF-B12 complex and another Cbl transport protein complex, trans-Cbl-B12, has been made. The pH effect on the binding of Cbl analogues in transport proteins is analyzed. A possible basis for the lack of interchangeability of human and rat IF receptors is presented.5] is a water-soluble vitamin and is essential for the growth and development of all mammals, including humans. Cbl is a member of the corrinoid series of cobalt-containing compounds and is distinguished from some other members of this group by possessing a nucleotide side chain terminating in dimethylbenzimidazole (1, 2). It serves as a key enzymatic cofactor for a number of methyltransferase and mutase reactions occurring in nature (3). In mammals, it contributes the prosthetic group for two important enzymes, methionine synthase (a methyltransferase) and methylmalonylCoA mutase (1, 2, 4).In mammals, three proteins are involved in the uptake, transport, and storage of Cbl. These are gastric intrinsic factor (IF), trans-Cbl II (TC), and haptocorrin (HC). These are immunologically distinct proteins with a protein core of Ϸ46 kDa. IF and HC are heavily glycosylated, but TC is not (5, 6). In humans, the genes for IF and HC are located in chromosome 22 (7,8), whereas the gene for TC is located in chromosome 11 (9). Human IF contains 399-aa residues plus Ϸ15% carbohydrate, giving it a molecular mass of Ϸ60 kDa, as observed by gel filtration (10-13). All three proteins promote Cbl entry through endocytosis involving distinct cell surface receptors (5,(14)(15)(16)(17)(18)(19)(20).Dietary Cbl is bound first to HC in the gastric lumen but is transferred to IF in the duodenum after degradation of HC by pancreatic proteases. IF is responsible for transit of Cbl through the small intestine and delivery of it to the endothelial cells that line the ileum. The IF-Cbl complex is then recognized by the IF-Cbl receptor, cubilin (CUB), located on the luminal side of the intestinal mucosal cells, and mediates its internalization. Lysosomal degradation of the internalized IF-Cbl complex releases Cbl, which is then able to bind to TC, probably within the enterocyte. The TC-Cbl complex is then translocated across the basolateral membrane and released into the circulation, to be taken up by TC-Cbl receptor-mediated process by all cells in the body. Lysosomal dissociation of the complex then occurs along with transfer of Cbl to the coenzyme methyl-and Ado-Cbl in the cytoplasm and i...

Structural Basis for Universal Corrinoid Recognition by the Cobalamin Transport Protein Haptocorrin

“…The ␣-domain consists of an intertwined ␣6/␣6 helical barrel as described for TC (20) and IF (21). However, conformational differences are observed at the ␣1-␣2, ␣3-␣4, and ␣5-␣6 loops, which are remote from the ligand interaction site.…”

“…1D and 2). In TC, no direct hydrogen bonds are observed at this interdomain contact, and in IF, only one salt bridge (Lys-365-Glu-110) and one hydrogen bond (Lys-365-Ser-105) have been described (21).…”

“…In the last 6 years, the crystal structures of human TC and bovine (bTC) TC (20), IF (21), and IF in complex with its receptor cubilin (22) have been reported. However, structural information on HC revealing the molecular determinants of the unique ability of HC to bind to Cbl analogs is still missing to date.…”

Trans and cis influences and effects in cobalamins and in their simple models

“…In other enzymes and in the transport proteins, the cobalamin binds to the protein in the so-called base-on form shown in Scheme 1 [37,38].…”

Vitamin B12 transport from food to the body's cells—a sophisticated, multistep pathway

“…Vitamin B 12 (B 12 [also known as cobalamin]; the abbrevia tion B 12 covers all forms of cobalamins and not only cyano cobalamin, which is the vitamin B 12 ) is a water soluble molecule that functions as an essential coenzyme for two enzymes in the human body: cytoplasmic methionine synthase, which catalyzes methylation of homo cysteine to methionine; and methylmalonylCoA mutase, which cata lyzes the conversion of methylmalonylCoA to succinyl CoA in the mitochondrion ( Figure 1). The methionine synthase reaction, which also involves folate (vitamin B 9 ) is essential for a high number of methyltransfer reactions and is also, therefore, indirectly involved in nucleotide synthesis.…”