Molecular determinants regulating Bruton’s tyrosine kinase activity and their mechanism: a combined computational and experimental approach

dc.contributor.advisor Amy H. Andreotti
dc.contributor.advisor Robert L. Jernigan Chopra, Nikita
dc.contributor.department Biochemistry, Biophysics and Molecular Biology 2018-08-11T06:36:20.000 2020-06-30T03:07:14Z 2020-06-30T03:07:14Z Fri Jan 01 00:00:00 UTC 2016 2017-08-31 2016-01-01
dc.description.abstract <p>Understanding allostery in proteins is critical in understanding their unique regulatory mechanisms and this knowledge can be exploited to develop highly specific, targeted therapies. In this dissertation, we have investigated the unique sequence elements that regulate the activity of a protein tyrosine kinase called Bruton’s tyrosine kinase or Btk. Btk is a member of the immune signaling pathway in B-cells and is required for B-cells maturation and function.</p> <p>Lack of a three-dimensional structure of full-length Btk kinase has proved a roadblock in understanding how the domains in Btk interact to shift its conformational equilibrium between active and inactive states. Moreover, in-spite of high homology between the catalytic centers of Btk and other well-studied protein tyrosine kinases such as Src, the regulatory mechanisms of these kinases appear to differ significantly creating an impediment to gaining a complete understanding of the mode of Btk regulation.</p> <p>In pursuit of the aim to identify key sequence and structure motifs that regulate Btk activity, we made use of a range of computational tools to better understand the Btk kinase domain and, when possible, the resulting hypotheses were tested using experimental methods. First, we have identified a specific isoleucine residue, conserved in Btk and related kinases, which functions to stabilize the inactive Btk conformation. We showed that substitution of the conserved isoleucine to leucine shifts the conformational equilibrium of the Btk kinase domain to the active state. Next, we showed how a highly conserved tryptophan, located in a linker region adjacent to the Btk kinase domain, stabilizes the active Btk kinase domain conformation through correlated dynamic motions within the kinase domain itself. Finally, sequence-structure information, combined with information theory and molecular dynamics, was used to identify a specific site in the Btk kinase domain that can be targeted to rescue the kinase activity of Btk in the presence of an inactivating disease causing mutation.</p> <p>The work presented here provides new insights into the regulatory mechanisms in Btk as well as potential allosteric sites in the protein, for which modulators of Btk activity could be developed. There is a growing need for the discovery of such allosteric modulators as Btk has been implicated in immunodeficiency disorders such as X-linked agammaglobulinemia as well as B-cells malignancies and breast and colon cancers. Ultimately, increased knowledge about the molecular mechanisms controlling Btk function should lead to the development of novel Btk activity modulators.</p>
dc.format.mimetype application/pdf
dc.identifier archive/
dc.identifier.articleid 6901
dc.identifier.contextkey 11169236
dc.identifier.s3bucket isulib-bepress-aws-west
dc.identifier.submissionpath etd/15894
dc.language.iso en
dc.source.bitstream archive/|||Fri Jan 14 20:48:02 UTC 2022
dc.subject.disciplines Biochemistry
dc.subject.disciplines Bioinformatics
dc.subject.keywords Bruton's Tyrosine Kinase
dc.subject.keywords Molecular Dynamics
dc.title Molecular determinants regulating Bruton’s tyrosine kinase activity and their mechanism: a combined computational and experimental approach
dc.type article
dc.type.genre dissertation
dspace.entity.type Publication
relation.isOrgUnitOfPublication faf0a6cb-16ca-421c-8f48-9fbbd7bc3747 Biochemistry dissertation Doctor of Philosophy
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