It is proposed to study the biological chemistry of sulfur in the thiol and disulfide forms and selenium in the selenol and diselenide forms. The long-term objectives of this research are to characterize chemical reactions which are central to the biological chemistry of sulfur and selenium, but which have not been quantitatively characterized because of a lack of experimental methods. Reactions will be characterized at the molecular level in intact erythrocytes, plasma and aqueous solution by NMR spectroscopy. Also, sensitive and selective methods based on high performance liquid chromatography (HPLC) with electrochemical detection will be developed and used to study the biological chemistry of thiols and disulfides in plasma and erythrocytes. The research on thiols and disulfides in intact erythrocytes will be designed to characterize intracellular thiol/disulfide chemistry under conditions of oxidative stress. Erythrocytes in vivo are regularly subjected to oxidative stress and conditions of oxidative stress exist in a diverse range of disease states. The proposed research will use noninvasive 1H-NMR methods to characterize the dynamics of intracellular thiol/disulfide chemistry of endogenous thiols and of thiol-containing drug molecules in erythrocytes as they are subjected to oxidative stress and as their intracellular region returns to its more normal reducing condition. HPLC methods will be used to determine disulfides formed at trace levels under these conditions, including hemoglobin-small molecule mixed disulfides. The chemistry of selenols and diselenides in intact erythrocytes will be studied by these same techniques with the objective of characterizing the intracellular chemistry of selenium in these forms. New and fundamental information about the oxidation-reduction chemistry of sulfur in amino acids, peptides, proteins and drug molecules will be obtained from studies of the kinetics and equilibria of thiol/disulfide exchange reactions in aqueous solution. The research will include studies of protein-small molecule mixed disulfides, which are through to be involved in processes as diverse as homocystinuria and drug-induced hypersensitivity, and studies of the disulfide groups in peptide hormones such as vasopressin and oxytocin. The kinetics and equilibria of reactions of thiols with diselenides and selenenylsulfides will also be characterized, with the objective of elucidating the chemistry of selenium at the active site of glutathione peroxidase.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Metallobiochemistry Study Section (BMT)
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University of California Riverside
Schools of Earth Sciences/Natur
United States
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Shi, Tiesheng; Spain, Stephen M; Rabenstein, Dallas L (2006) A striking periodicity of the cis/trans isomerization of proline imide bonds in cyclic disulfide-bridged peptides. Angew Chem Int Ed Engl 45:1780-3
Shi, Tiesheng; Rabenstein, Dallas L (2002) Convenient synthesis of human calcitonin and its methionine sulfoxide derivative. Bioorg Med Chem Lett 12:2237-40
Kaerner, A; Rabenstein, D L (1999) Stability and structure-forming properties of the two disulfide bonds of alpha-conotoxin GI. Biochemistry 38:5459-70
Huang, H; Rabenstein, D L (1999) A cleavage cocktail for methionine-containing peptides. J Pept Res 53:548-53
Rabenstein, D L; Russell, J; Gu, J (1998) Characterization of the thiol/disulfide chemistry of peptides corresponding to the 603-609 disulfide loop of the human immunodeficiency virus (HIV) envelope glycoprotein gp41. J Pept Res 51:437-43
Rabenstein, D L; Hari, S P; Kaerner, A (1997) Determination of acid dissociation constants of peptide side-chain functional groups by two-dimensional NMR. Anal Chem 69:4310-6
Russell, J; Rabenstein, D L (1996) Speciation and quantitation of underivatized and Ellman's derivatized biological thiols and disulfides by capillary electrophoresis. Anal Biochem 242:136-44
Rabenstein, D L; Millis, K K (1995) Nuclear magnetic resonance study of the thioltransferase-catalyzed glutathione/glutathione disulfide interchange reaction. Biochim Biophys Acta 1249:29-36
Yeo, P L; Rabenstein, D L (1993) Characterization of the thiol/disulfide chemistry of neurohypophyseal peptide hormones by high-performance liquid chromatography. Anal Chem 65:3061-6
Keire, D A; Mariappan, S V; Peng, J et al. (1993) Nuclear magnetic resonance studies of the binding of captopril and penicillamine by serum albumin. Biochem Pharmacol 46:1059-69

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