Elektrophysiologische Charakterisierung des mitochondrialen Porins VDAC1 und des antimikrobiellen Peptids Dermcidin in lösungsmittelfreien Modellmembranen

Weichbrodt, Conrad

dc.contributor.advisor	Steinem, Claudia Prof. Dr.	de
dc.contributor.author	Weichbrodt, Conrad	de
dc.date.accessioned	2013-07-04T09:08:23Z	de
dc.date.available	2013-07-04T09:08:23Z	de
dc.date.issued	2013-07-04	de
dc.identifier.uri	http://hdl.handle.net/11858/00-1735-0000-0001-BAA4-C	de
dc.identifier.uri	http://dx.doi.org/10.53846/goediss-3921
dc.language.iso	deu	de
dc.publisher	Niedersächsische Staats- und Universitätsbibliothek Göttingen	de
dc.rights.uri	http://creativecommons.org/licenses/by-nc-nd/3.0/
dc.subject.ddc	540	de
dc.title	Elektrophysiologische Charakterisierung des mitochondrialen Porins VDAC1 und des antimikrobiellen Peptids Dermcidin in lösungsmittelfreien Modellmembranen	de
dc.type	doctoralThesis	de
dc.title.translated	Electrophysilogical characterization of the mitochondrial porin VDAC1 and the antimicrobial peptide Dermcidin in solvent-free model membranes	de
dc.contributor.referee	Steinem, Claudia Prof. Dr.	de
dc.date.examination	2013-04-12	de
dc.description.abstracteng	Solvent free lipid bilayers were used for the electrophysiological characterization of the most abundant protein of the outer mitochondrial membrane, the isoform 1 of the voltage dependent anion channel (VDAC1), and the antimicrobial peptide Dermcidin. The bilayers were prepared via spreading of giant unilamellar vesicles (GUVs) over a single pore in a borosilicate glass surface. To gain deeper insight in its gating behavior, wild type VDAC1 as well as two VDAC1 mutants were incorporated into GUVs which then were spread over the aperture. Subsequent analysis of single channels at varying transmembrane potentials Um revealed the transition between an open conformation with a conductance of Go = 4.0 nS at low potentials and a closed conformation with Gc = 2.0 nS in 1 M KCl for Um ≥ 30 mV and Um ≤ 30 mV for all VDAC1 variants. This gating between the main VDAC1 states with low transition rates of kmg = 0.1 2 s-1 did not necessarily occur at high transmembrane potentials as the open probability Po did not fall below 90 % even at Um ≥ 30 mV on a single channel level. Furthermore, a VDAC1 mutant with three additional amino acids at its N-terminal end (RGS VDAC1) exhibited a previously not characterized second gating that superimposed the main gating of VDAC1. Closer analysis of this fast gating resulted in a lower amplitude of ΔGfg = 0.5 1.2 nS and much higher transition rates of kfg = 18 911 s 1 with higher rates at low voltages and vice versa. A second mutant (V17C/A205C VDAC1) contained an artificially introduced disulfide bond that prevented the movement of a certain protein structure as putative molecular basis of the VDAC1 conductance transition. However, V17C/A205C VDAC1 did not exhibit an altered behavior compared to the wild type VDAC1. The antimicrobial activity of Dermcidin (DCD) mainly consists of its ability to perforate lipid bilayers. To elucidate the postulated zinc dependency of the DCD activity, stable membranes were prepared and subsequently incubated with DCD in a zinc free environment or in the presence of Zn2+ ions. High DCD activity was detected in the presence of Zn2+ ions whereas no activity was found under zinc free conditions or in the presence of Mg2+ ions. These findings and the inactivation of DCD via the alteration of a zinc binding site in a mutant (H38A DCD) proved the zinc dependency of the DCD activity.	de
dc.contributor.coReferee	Schroeder, Jörg Prof. Dr.	de
dc.contributor.thirdReferee	Diederichsen, Ulf Prof. Dr.	de
dc.contributor.thirdReferee	Ficner, Ralf Prof. Dr.	de
dc.contributor.thirdReferee	Tittmann, Kai Prof. Dr.	de
dc.contributor.thirdReferee	Meinecke, Michael Dr.	de
dc.subject.ger	Elektrophysiologie	de
dc.subject.ger	Ionenkanal	de
dc.subject.ger	antimikrobielles Peptid	de
dc.subject.ger	Modellmembranen	de
dc.subject.ger	Dermcidin	de
dc.subject.ger	VDAC1	de
dc.subject.eng	ionchannels	de
dc.subject.eng	electrophysiology	de
dc.subject.eng	antimicrobial peptides	de
dc.subject.eng	VDAC1	de
dc.subject.eng	Dermcidin	de
dc.subject.eng	model membranes	de
dc.identifier.urn	urn:nbn:de:gbv:7-11858/00-1735-0000-0001-BAA4-C-9	de
dc.affiliation.institute	Fakultät für Chemie	de
dc.subject.gokfull	Chemie (PPN62138352X)	de
dc.identifier.ppn	751154482	de

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