Abstract

Vitamin B12 (cobalamin) deficiency in man leads to two broad classes of clinical phenomena: 1) megaloblastic anemia and related changes in rapidly proliferating tissues and, 2) a variety of neurological disorders. The biochemical basis of these chemical manifestations is not at all understood and indeed the mechanism of the cobalamin-dependent enzymatic reactions has not yet been elucidated. Our proposal outlines experiments that are designed to provide information about the reactivity of the carbon-cobalt bond of the corrinoid coenzymes. What interaction with the enzyme causes the reversible homolysis of this organometalic bond? How does the enzyme prevent the very reactive radical intermediates from spurious reactions? We have selected the adenosylcobalamin-dependent ribonucleotide reductase system as the target enzyme system. The proposed studies are directed towards: a. The characterization of the active site of the ribonucleotide reductase from Corynebacterium nephridii. b. The synthesis and evaluation of possible transition state analogs that will inhibit ribonucleotide reduction and thus cell proliferation. c. 13C-NMR studies of thioredoxin, modified with [13C]methyl iodide and [13C]formaldehyde. d. The purification and characterization of thioredoxin reductase from C. nephridii.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM033776-02
Application #
3283755
Study Section
Biochemistry Study Section (BIO)
Project Start
1984-07-01
Project End
1989-06-30
Budget Start
1985-07-01
Budget End
1986-06-30
Support Year
2
Fiscal Year
1985
Total Cost
Indirect Cost

  Institution

Name
University of Minnesota Twin Cities
Department
Type
Schools of Medicine
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455

  Publications

Krone, U E; McFarlan, S C; Hogenkamp, H P (1994) Purification and partial characterization of a putative thymidylate synthase from Methanobacterium thermoautotrophicum. Eur J Biochem 220:789-94
Loida, P J; Sligar, S G (1993) Molecular recognition in cytochrome P-450: mechanism for the control of uncoupling reactions. Biochemistry 32:11530-8
Ong, S P; McFarlan, S C; Hogenkamp, H P (1993) 2'-C-methyladenosine and 2'-C-methyluridine 5'-diphosphates are mechanism-based inhibitors of ribonucleoside diphosphate reductase from Corynebacterium nephridii. Biochemistry 32:11397-404
Ong, S P; Nelson, L S; Hogenkamp, H P (1992) Synthesis of 3'-C-methyladenosine and 3'-C-methyluridine diphosphates and their interaction with the ribonucleoside diphosphate reductase from Corynebacterium nephridii. Biochemistry 31:11210-5
Schallreuter, K U; Elgren, T E; Nelson Jr, L S et al. (1992) Ribonucleotide diphosphate reductase from human metastatic melanoma. Melanoma Res 2:393-400
McFarlan, S C; Terrell, C A; Hogenkamp, H P (1992) The purification, characterization, and primary structure of a small redox protein from Methanobacterium thermoautotrophicum, an archaebacterium. J Biol Chem 267:10561-9
Sze, I S; McFarlan, S C; Spormann, A et al. (1992) A possible new class of ribonucleotide reductase from Methanobacterium thermoautotrophicum. Biochem Biophys Res Commun 184:1101-7
McFarlan, S C; Hogenkamp, H P; Eccleston, E D et al. (1989) Purification, characterization and revised amino acid sequence of a second thioredoxin from Corynebacterium nephridii. Eur J Biochem 179:389-98
Mirkin, B L; O'Dea, R F; Hogenkamp, H P (1987) Cytotoxic action of adenosine nucleoside and dialdehyde analogues on murine neuroblastoma in tissue culture: structure-activity relationships. Cancer Res 47:3650-5
Lim, C J; Fuchs, J A; McFarlan, S C et al. (1987) Cloning, expression, and nucleotide sequence of a gene encoding a second thioredoxin from Corynebacterium nephridii. J Biol Chem 262:12114-9

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