Specific enzymic interactions relating to the mechanism of action on certain 2-chloroethyl nitrosoureas Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/s4655k631

Descriptions

Attribute NameValues
Creator
Abstract or Summary
  • Certain 2- chioroethyl nitrosoureas have been demonstrated to be highly specific inactivators of chymotrypsin in vitro and glutathione reductase, both in vivo and in isolated rat hepatocytes. The inactivation of glutathione reductase in isolated hepatocytes resulted in an increased sensitivity toward an adriamycin induced oxidative challenge. Prolonged incubation of 1-(2-chloroethyl)-3-([1-¹⁴ C] cyclohexyl)-1-nitrosourea (CCNU) with chymotrypsin resulted in covalent modification and concomitant inactivation of chymotrypsin via degradation of the nitrosourea to form cyclohexyl isocyanate. Cyclohexyl isocyanate was shown to be an active-site-directed inactivator of chymotrypsin. L-Lysine did not protect the enzyme from inactivation by cyclohexyl isocyanate. Degradation of an excess of 1-(2-chloroethyl)-3- ([1-¹⁴c] -cyclohexyl)-1-nitrosourea in the presence of*enzyme yielded 1.11 ± .07 mole of enzyme inactivated. Short-term incubation demonstrated that the nitrosourea neither inhibited nor protected the enzyme from cyclohexyl isocyanate inactivation. Treatment of chymotrypsin with less than stoichiometric amounts of cyclohexyl isocyanate or titration of the active site serine with phenylmethanesulfonylfluoride prior to [ ¹⁴ C]- labeling resulted in a proportional decrease in bound [ ¹⁴C] cyclohexyl moiety. In addition, the specific inactivation of yeast glutathione reductase by 2-chloroethyl isocyanate and cyclohexyl isocyanate derived from their respective 2-chloroethyl nitrosoureas has been demonstrated. Titration of the enzyme with 2-chloroethyl isocyanate or [ ¹⁴ C] labeling with 1-(2-chloroethyl)-3-([1- ¹⁴C]chclohexyl)-1-nitrosourea or 1,3-bis-(2-[¹⁴C]chloroethyl)-1-nitrosourea (BCNU) resulted in a near stoichiometric inactivation and/or covalent labeling of the enzyme. In addition to BCNU and CCNU several other 2-chloroethyl nitrosoureas were capable of inactivation of not only purified glutathione reductase, but also the activity of this enzyme in cell-free extracts of murine lymphoma L5178Y ascites tumor cells and murine bone marrow. A positive correlation has been shown between the ability of certain 2-chloroethyl nitrosoureas to inactivate glutathione reductase and the degree of myelotoxicity observed for these nitrosoureas. Incubation of isolated rat hepatocytes with BCNU resulted in the selective and extensive (>90%) inactivation of hepatic glutathione reductase. BCNU also depleted intracellular glutathione by 70% but had no significant effect on cell viability or lipid peroxidation. Incubation of BCNU-treated hepatocytes with adriamycin (ADR) resulted in a decrease in cell viability concurrent with an increase in lipid peroxidation. These effects were not observed with untreated hepatocytes incubated with ADR. Glutathione depletion with diethyl maleate and incubation with ADR did not result in increased cellular damage or lipid peroxidation. Incubation of BCNU-treated hepatocytes with ADR in the presence of a-tocopherol resulted in a significant amount of protection from ADR damage.
Resource Type
Date Available
Date Copyright
Date Issued
Degree Level
Degree Name
Degree Field
Degree Grantor
Commencement Year
Advisor
Academic Affiliation
Non-Academic Affiliation
Subject
Rights Statement
Peer Reviewed
Language
Digitization Specifications
  • File scanned at 300 ppi (Monochrome) using ScandAll PRO 1.8.1 on a Fi-6770A in PDF format. CVista PdfCompressor 5.0 was used for pdf compression and textual OCR.
Replaces
Additional Information
  • description.provenance : Submitted by Kirsten Clark (kcscannerosu@gmail.com) on 2013-08-22T23:11:10Z No. of bitstreams: 1 BabsonJohnRobert1981.pdf: 737849 bytes, checksum: 4e0492c922b6f39d9243299cb522ae61 (MD5)
  • description.provenance : Approved for entry into archive by Patricia Black(patricia.black@oregonstate.edu) on 2013-08-27T17:30:31Z (GMT) No. of bitstreams: 1 BabsonJohnRobert1981.pdf: 737849 bytes, checksum: 4e0492c922b6f39d9243299cb522ae61 (MD5)
  • description.provenance : Approved for entry into archive by Patricia Black(patricia.black@oregonstate.edu) on 2013-08-23T18:31:14Z (GMT) No. of bitstreams: 1 BabsonJohnRobert1981.pdf: 737849 bytes, checksum: 4e0492c922b6f39d9243299cb522ae61 (MD5)
  • description.provenance : Made available in DSpace on 2013-08-27T17:30:31Z (GMT). No. of bitstreams: 1 BabsonJohnRobert1981.pdf: 737849 bytes, checksum: 4e0492c922b6f39d9243299cb522ae61 (MD5) Previous issue date: 1980-08-29

Relationships

In Administrative Set:
Last modified: 08/17/2017

Downloadable Content

Download PDF
Citations:

EndNote | Zotero | Mendeley

Items