Mechanistic investigations of novel, irreversible inhibitors of S-adenosylhomocysteine hydrolase
| dc.contributor.advisor | Parry, Ronald J. | |
| dc.creator | Muscate, Angelika | |
| dc.date.accessioned | 2009-06-04T00:03:22Z | |
| dc.date.available | 2009-06-04T00:03:22Z | |
| dc.date.issued | 1992 | |
| dc.description.abstract | Isolation of bovine S-adenosylhomocysteine hydrolase by affinity chromatography unexpectedly yielded two forms of the enzyme. Type A contained 4 moles of NAD$\sp+$/mole of enzyme tetramer, while Type B had only half that number of nucleotide cofactors associated with it. The acetylenic analog of adenosine, 9-(5$\sp\prime$,6$\sp\prime$-dideoxy-$\beta$-D-ribo-hex-5$\sp\prime$-ynofuranosyl)-adenine, was synthesized, and its behavior as an inhibitor of Type A enzyme examined. Irreversible inactivation of the enzyme with excess inhibitor was accompanied by the reduction of 2 equivalents of NAD$\sp+$ to NADH, release of the remaining NAD$\sp+$ from the enzyme, and binding of 4 equivalents of inhibitor per enzyme tetramer. Denaturation studies suggested that two equivalents of the inhibitor may form a covalent bond between the enzyme and the oxidized inhibitor. The inactivation behavior exhibited by Type A and Type B enzyme with 2$\sp\prime$-deoxy-2$\sp\prime$,2$\sp\prime$-difluoroadenosine was compared. The Type B enzyme was irreversibly inactivated by excess inhibitor with concurrent reduction of 1 mole of NAD$\sp+$ to NADH and release of 1 mole of NAD$\sp+$. Inactivation of the Type A enzyme was accompanied by the reduction of 1 mole of NAD$\sp+$ to NADH and release of ca. 2 moles of NAD$\sp+$. Upon dialysis, 50% of the original activity of the enzyme was regained. Binding stoichiometries of 1 and 3 moles inhibitor/mole of enzyme tetramer were determined for the Type B and Type A enzyme, respectively. Denaturation studies indicated that 1 equivalent of the oxidized inhibitor may have formed a covalent bond with the Type A enzyme, but not with Type B. The binding behavior of adenosine with both types of the enzyme was also studied. Type B enzyme exhibited a large binding affinity of 4-25 moles of adenosine/mole of enzyme tetramer, while Type A bound only 3 moles of adenosine/mole enzyme tetramer. The binding of adenosine to the Type B enzyme was accompanied by the reduction of 1 equivalent of NAD$\sp+$ to NADH. Denaturation experiments suggested the formation of a covalent bond between the enzyme and one equivalent of adenosine. However, peptide mapping of the reduced enzyme-adenosine complex failed. | |
| dc.format.extent | 127 p. | en_US |
| dc.format.mimetype | application/pdf | |
| dc.identifier.callno | Thesis Chem. 1992 Muscate | |
| dc.identifier.citation | Muscate, Angelika. "Mechanistic investigations of novel, irreversible inhibitors of S-adenosylhomocysteine hydrolase." (1992) Diss., Rice University. <a href="https://hdl.handle.net/1911/16532">https://hdl.handle.net/1911/16532</a>. | |
| dc.identifier.uri | https://hdl.handle.net/1911/16532 | |
| dc.language.iso | eng | |
| dc.rights | Copyright is held by the author, unless otherwise indicated. Permission to reuse, publish, or reproduce the work beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder. | |
| dc.subject | Organic chemistry | |
| dc.subject | Biochemistry | |
| dc.title | Mechanistic investigations of novel, irreversible inhibitors of S-adenosylhomocysteine hydrolase | |
| dc.type | Thesis | |
| dc.type.material | Text | |
| thesis.degree.department | Chemistry | |
| thesis.degree.discipline | Natural Sciences | |
| thesis.degree.grantor | Rice University | |
| thesis.degree.level | Doctoral | |
| thesis.degree.name | Doctor of Philosophy |
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