The long term objectives of this project are to characterize precisely the interaction of cobalamin with macromolecules required for intestinal absorption, transport, and cellular uptake of this vitamin and to develop more readily applicable radioassays by which to identify, measure, and study the metabolism of the physiologic and non-physiologic cobalamins. Toward these objectives, we have the following specific aims: a) To characterize the mechanism by which the intrinsic factor (IF)-cobalamin complex binds to the IF-receptor protein and facilitates the internalization of cobalamin. For this we will use radioimmunoassay and immunocytochemical analysis of ileal epithelial cells. b) To identify and characterize immunoreactive forms of the cobalamin binding proteins fractionated by gel-filtration and ion exchange chromatography using specific RIA to identify these proteins. c) To develop radioimmunoassay techniques to measure the specific physiologic forms of cobalamin and use these analytical methods to measure the concentration of these coenzymes directly in blood and tissue extracts. d) To isolate and characterize the nonphysiologic forms of corrinoid compounds in plasma using chromatographic separation and ligand binding radioassays. e) To develop radioimmunoenzymatic assays to study the synthesis of the two physiologic coenzymes, adenosyl cobalamin and methyl cobalamin and to characterize the enzymes required for these biosynthetic processes. The information obtained and the techniques developed in this project would extend our understanding of the disorders of cobalamin absorption and transport which may occur with drugs, toxins and liver disease; and the analytical techniques to measure cobalamin coenzymes could provide a far simpler method for determining acquired and inherited cellular disorders of cobalamin coenzyme synthesis.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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General Medicine A Subcommittee 2 (GMA)
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Suny Downstate Medical Center
Schools of Medicine
United States
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Qian, Lian; Quadros, Edward V; Regec, Annette et al. (2002) Congenital transcobalamin II deficiency due to errors in RNA editing. Blood Cells Mol Dis 28:134-42; discussion 143-5
Sobti, P; Rothenberg, S P; Quadros, E V (2000) Radioenzymatic assay for reductive catalysis of N(5)N(10)-methylenetetrahydrofolate by methylenetetrahydrofolate reductase. J Biochem Biophys Methods 46:20-Nov
Quadros, E V; Regec, A L; Khan, K M et al. (1999) Transcobalamin II synthesized in the intestinal villi facilitates transfer of cobalamin to the portal blood. Am J Physiol 277:G161-6
Wilbur, D S; Pathare, P M; Hamlin, D K et al. (1999) Radioiodination of cyanocobalamin conjugates containing hydrophilic linkers: preparation of a radioiodinated cyanocobalamin monomer and two dimers, and assessment of their binding with transcobalamin II. Bioconjug Chem 10:912-20
Rothenberg, S P (1999) Increasing the dietary intake of folate: pros and cons. Semin Hematol 36:65-74
McLean, G R; Quadros, E V; Rothenberg, S P et al. (1997) Antibodies to transcobalamin II block in vitro proliferation of leukemic cells. Blood 89:235-42
Quadros, E V; Rothenberg, S P; McLoughlin, P (1996) Characterization of monoclonal antibodies to epitopes of human transcobalamin II. Biochem Biophys Res Commun 222:149-54
Rothenberg, S P; Quadros, E V (1995) Transcobalamin II and the membrane receptor for the transcobalamin II-cobalamin complex. Baillieres Clin Haematol 8:499-514
Regec, A; Quadros, E V; Platica, O et al. (1995) The cloning and characterization of the human transcobalamin II gene. Blood 85:2711-9
Quadros, E V; Sai, P; Rothenberg, S P (1994) Characterization of the human placental membrane receptor for transcobalamin II-cobalamin. Arch Biochem Biophys 308:192-9

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