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dc.contributor.advisor Görlich, Dirk Prof. Dr.
dc.contributor.author Gencalp, Kevser
dc.date.accessioned 2015-08-18T10:24:46Z
dc.date.available 2015-08-18T10:24:46Z
dc.date.issued 2015-08-18
dc.identifier.uri http://hdl.handle.net/11858/00-1735-0000-0022-6082-0
dc.language.iso eng de
dc.relation.uri http://creativecommons.org/licenses/by-nc-nd/4.0/
dc.subject.ddc 570 de
dc.title Nuclear export of actin: A biochemical and structural perspective de
dc.type doctoralThesis de
dc.contributor.referee Görlich, Dirk Prof. Dr.
dc.date.examination 2014-10-24
dc.description.abstracteng Macromolecular exchange between the nucleus and the cytoplasm is mainly mediated by the RanGTP-dependent nuclear transport receptors (NTRs) of the Impβ family. NTRs are classified as importins and exportins according to the direction of transport. Exportins carry their cargos from the nucleus to cytoplasm as a ternary export complex together with RanGTP. Impβ-like NTRs are made up of multiple HEAT repeats and share an inherently flexible structure. Nevertheless, structural analyses so far revealed unique cargo recognition mechanisms for each NTR, and even for different cargos recognized by the same receptor. Actin is the well-known constituent of the cytoplasmic microfilaments. Monomeric actin with its small size and globular fold can slowly diffuse into the nucleus. Exportin 6 (Xpo6) is a dedicated export receptor found in vertebrates and insects, whose only known function is to export nuclear actinprofilin back to cytoplasm. In the absence of Xpo6, actin accumulates in the nucleus. Amphibian oocytes exploit this to stabilize their giant nuclei with an intranuclear actin cytoskeleton by blocking Xpo6 expression. On the other hand, nuclear accumulation of actin is also seen in intranuclear rod myopathies (IRM), devastating muscle diseases caused by several mutations of the skeletal alpha actin gene. We hypothesize that these mutations of actin interfere with its recognition and nuclear export by Xpo6, resulting in the characteristic nuclear accumulations. To decipher the cargo recognition by Xpo6 and to understand the mechanisms underlying the IRM, we set out to crystallize the Xpo6*RanGTP*actin*profilin complex. It turned out that the actin export complex is extremely salt sensitive, and muscle and non-muscle actin isoforms differ in their affinity for profilin and hence, for the export complex. We developed a new single-step protocol for purification of profilin*β/γ-actin complexes from cytoplasmic extracts, which enabled us to purify a stable actin export complex. Topological analysis of the actin export complex showed that the Poly-Proline-binding pocket of profilin is accessible in the complex, whereas the binding site for DNaseI on actin overlaps with the Xpo6 binding site. Crystals of cargo-free Xpo6 were obtained in several different conditions. However, the diffraction is currently limited to 7.4 Å resolution. Also the actin nuclear export complex was successfully crystallized. These crystals will be the substrates for future structural analyses and will help us to understand the cargo recognition by Xpo6. de
dc.contributor.coReferee Großhans, Jörg Prof. Dr.
dc.contributor.thirdReferee Rehling, Peter Prof. Dr.
dc.contributor.thirdReferee Stark, Holger Prof. Dr.
dc.contributor.thirdReferee Doenecke, Detlef Prof. Dr.
dc.subject.ger actin de
dc.subject.ger nuclear export de
dc.subject.ger exportin 6 de
dc.subject.ger profilin de
dc.subject.eng actin de
dc.subject.eng nuclear export de
dc.subject.eng exportin 6 de
dc.subject.eng profilin de
dc.identifier.urn urn:nbn:de:gbv:7-11858/00-1735-0000-0022-6082-0-2
dc.affiliation.institute Göttinger Graduiertenschule für Neurowissenschaften, Biophysik und molekulare Biowissenschaften (GGNB) de
dc.subject.gokfull Biologie (PPN619462639) de
dc.identifier.ppn 833463640

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