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dc.contributor.authorRudyak, Stanislav Gennadyevich
dc.date.accessioned2018-07-12T17:02:06Z
dc.date.available2018-07-12T17:02:06Z
dc.date.issued2001
dc.identifier.citationSource: Dissertation Abstracts International, Volume: 62-04, Section: B, page: 1859.;Advisors: Thomas E. Shrader.
dc.identifier.urihttps://yulib002.mc.yu.edu/login?url=http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqm&rft_dat=xri:pqdiss:3012327
dc.identifier.urihttps://hdl.handle.net/20.500.12202/520
dc.description.abstractProtein degradation mediates a variety of biological processes ranging from regulation of the some metabolic pathways to housekeeping. In prokaryotes, ATP-dependent proteases mediate the turnover of abnormal and damaged proteins and degradation of some specific short-lived proteins. The Lon (La) protease is responsible for degradation of the majority of abnormal proteins synthesized in Escherichia coli. It is a multimeric enzyme that is activated by ATP and Mg2+ ions, and stimulated by unfolded proteins. In recent years homologs of Lon protease have been discovered in many prokaryotic organisms and in mitochondria. The Lon protease from Mycobacterium smegmatis (Ms-Lon) was selected as a model to study the ATP-dependent protein degradation by Lon. Results show that Ms-Lon can interact with three proteolytic substrates simultaneously and that formation of this quaternary complex requires the N-terminal region of Ms-Lon. Overall, our data support models of Ms-Lon that include two allosteric polypeptide binding sites distinct from the catalytic peptidase site. To characterize the structural dependence of substrate recognition by Ms-Lon, a series of mutants of the nuclease from Staphylococcus aureus (SNase) were tested as substrates of Ms-Lon. Degradation of these mutants was strongly correlated with their overall degree of compaction. In addition, the substrates containing low levels of secondary structure required both ATP hydrolysis and an intact Ms-Lon's N-terminal domain for degradation. We conclude that both sequence and structural components are important for substrate recognition by Ms-Lon. Finally, the peptidase activity of Ms-Lon was dependent upon both its concentration and that of Mg2+, suggesting that Mg 2+-dependent oligomerization modulates the peptidase activity of Ms-Lon. Indeed, analytical ultracentrifugation data demonstrated that self-association of Ms-Lon was enhanced in the presence of Mg2+. Subsequently, a stimulatory effect of alpha-casein on Ms-Lon's self-association was suggested by the chemical dissociation experiments. These results show that activation of Ms-Lon requires oligomerization and that Ms-Lon self-association is facilitated by its activator, Mg2+, and stimulator, unfolded protein. (Abstract shortened by UMI.).
dc.publisherProQuest Dissertations & Theses
dc.subjectBiochemistry.
dc.titleMechanistic studies of the Lon protease from Mycobacterium smegmatis
dc.typeDissertation


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