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dc.contributor.advisorRobert T. Sauer and Tania A. Baker.en_US
dc.contributor.authorStinson, Benjamin M. (Benjamin Michael)en_US
dc.contributor.otherMassachusetts Institute of Technology. Department of Biology.en_US
dc.date.accessioned2015-06-10T18:42:07Z
dc.date.available2015-06-10T18:42:07Z
dc.date.copyright2014en_US
dc.date.issued2015en_US
dc.identifier.urihttp://hdl.handle.net/1721.1/97273
dc.descriptionThesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, February 2015.en_US
dc.descriptionThis electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.en_US
dc.descriptionCataloged from student-submitted PDF version of thesis.en_US
dc.descriptionIncludes bibliographical references.en_US
dc.description.abstractATP-powered proteases enforce protein quality-control and regulation in all domains of life. ClpX, a AAA+ ring homohexamer, uses the energy of ATP binding and hydrolysis to power conformational changes that unfold and translocate target proteins into the ClpP peptidase for degradation. X-ray crystal structures show that some ClpX subunits occupy nucleotide-loadable conformations and others occupy unloadable conformations. Furthermore, biochemical evidence suggests that multiple classes of nucleotide-loadable subunits exist. How asymmetry among subunits is coordinated to achieve mechanical function has remained unclear. Using a combination of mutagenesis, disulfide crosslinking, and fluorescence methods to assay the conformations and nucleotide-binding properties of individual subunits, we demonstrate dynamic interconversion of loadable and unloadable subunits. Such interconversion is required to couple ATP hydrolysis by ClpX to mechanical work, plays a role in substrate binding and ClpP interaction, and is not strictly coupled to the ATP hydrolysis cycle. ATP binding to different classes of subunits drives allosteric changes in ring conformation to allow hydrolysis and coupled machine function, and we present a subunit-specific single molecule nucleotide occupancy assay to elucidate details of this process.en_US
dc.description.statementofresponsibilityby Benjamin M. Stinson.en_US
dc.format.extent140 pagesen_US
dc.language.isoengen_US
dc.publisherMassachusetts Institute of Technologyen_US
dc.rightsM.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.en_US
dc.rights.urihttp://dspace.mit.edu/handle/1721.1/7582en_US
dc.subjectBiology.en_US
dc.titleNucleotide binding and conformational switching in the hexameric ring of a AAA+ machineen_US
dc.typeThesisen_US
dc.description.degreePh. D.en_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Biology.en_US
dc.identifier.oclc910720311en_US


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