Abstract
In humans, three soluble extracellular cobalamin-binding proteins; transcobalamin (TC), intrinsic factor (IF), and haptocorrin (HC), are involved in the uptake and transport of cobalamin. In this study, we investigate a cobalamin-binding protein from zebrafish (Danio rerio) and summarize current knowledge concerning the phylogenetic evolution of kindred proteins. We identified a cobalamin binding capacity in zebrafish protein extracts (8.2 pmol/fish) and ambient water (13.5 pmol/fish) associated with a single protein. The protein showed resistance toward degradation by trypsin and chymotrypsin (like human IF, but unlike human HC and TC). The cobalamin analogue, cobinamide, bound weaker to the zebrafish cobalamin binder than to human HC, but stronger than to human TC and IF. Affinity for another analogue, adenosyl-pseudo-cobalamin was low compared with human HC and TC, but high compared with human IF. The absorbance spectrum of the purified protein in complex with hydroxo-cobalamin resembled those of human HC and IF, but not TC. We searched available databases to further explore the phylogenies of the three cobalamin-binding proteins in higher vertebrates. Apparently, TC-like proteins are the oldest evolutionary derivatives followed by IF and HC (the latter being present only in reptiles and most but not all mammals). Our findings suggest that the only cobalamin-binding protein in zebrafish is an intermediate between the three human cobalamin binders. These findings support the hypothesis about a common ancestral gene for all cobalamin-binding proteins in higher vertebrates.
Highlights
The uptake and transport of cobalamin is mediated by three structural kindred proteins: transcobalamin (TC) that transports cobalamin through the bloodstream and ensures its cellular uptake; haptocorrin (HC) that carries most of the circulating cobalamin as well as inactive analogues of the vitamin; and intrinsic factor (IF) that facilitates the intestinal uptake of the vitamin [1,2]
We investigated the soluble cobalaminbinding protein from zebrafish (Danio rerio) and compared it with the three known soluble extracellular cobalamin-binding proteins present in humans and other mammals
Those are IF, the protein facilitating intestinal uptake of cobalamin; TC, the protein mediating the uptake of cobalamin by the cells of the body; and HC, a protein of unknown function present in plasma and most extracellular fluids [1,2]
Summary
The uptake and transport of cobalamin (vitamin B12) is mediated by three structural kindred proteins: transcobalamin (TC) that transports cobalamin through the bloodstream and ensures its cellular uptake; haptocorrin (HC) that carries most of the circulating cobalamin as well as inactive analogues of the vitamin; and intrinsic factor (IF) that facilitates the intestinal uptake of the vitamin [1,2]. The cobalamin-binding proteins seemingly evolved from a common ancestral gene [3,4,6], where sequential duplication of genes caused divergence of IF from TC and later on HC from IF [1]. The phylogenetic search for cobalamin binders within the subphylum vertebrata has mostly been restricted to mammals. The proteins TC and IF (as well as their encoding genes, TCN2 and GIF, respectively) have been found in all investigated species, including mouse, rat, hog, cow, and chimpanzee [10]. HC and/or its encoding gene, TCN1, have been found in many mammals including hog [6], rabbit [11], and cow [12]. Recent studies show that mice lack HC, and that TC is their only cobalamin transporter in blood resembling both human TC and HC [13]
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