comparison of recombinant human haptocorrin expressed in human embryonic kidney cells and native haptocorrin比较重组人类haptocorrin表达人类胚胎肾细胞和原生haptocorrin.pdf

Comparison of Recombinant Human Haptocorrin Expressed in Human Embryonic Kidney Cells and Native Haptocorrin 1 2 3 1 1 Evelyne Furger , Sergey N. Fedosov , Dorte Launholt Lildballe , Robert Waibel , Roger Schibli , 3 1 Ebba Nexo , Eliane Fischer * 1 Center for Radiopharmaceutical Sciences, Paul Scherrer Institute, Villigen, Switzerland, 2 Department of Engineering, Aarhus University, Aarhus, Denmark, 3 Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark Abstract Haptocorrin (HC) is a circulating corrinoid binding protein with unclear function. In contrast to transcobalamin, the other transport protein in blood, HC is heavily glycosylated and binds a variety of cobalamin (Cbl) analogues. HC is present not only in blood but also in various secretions like milk, tears and saliva. No recombinant form of HC has been described so far. We report the expression of recombinant human HC (rhHC) in human embryonic kidney cells. We purified the protein with a yield of 6 mg (90 nmol) per litre of cell culture supernatant. The isolated rhHC behaved as native HC concerning its spectral properties and ability to recognize both Cbl and its baseless analogue cobinamide. Similar to native HC isolated from blood, rhHC bound to the asialoglycoprotein receptor only after removal of terminal sialic acid residues by treatment with neuraminidase. Interestingly, rhHC, that compared to native HC contains four excessive amino acids (…LVPR) at the C- terminus, showed subtle changes in the binding kinetics of Cbl, cobinamide and the fluorescent Cbl conjugate CBC. The recombinant protein has properties very similar to native HC and although showing slightly different