Molecular Details of Membrane Deformation by ENTH Domains
by Benjamin Kroppen
Date of Examination:2017-11-22
Date of issue:2018-03-20
Advisor:Prof. Dr. Michael Meinecke
Referee:Prof. Dr. Andreas Janshoff
Referee:Dr. Achim Dickmanns
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Abstract
English
During clathrin mediated endocytosis (CME) cargo molecules are transported across the plasma membrane from the outer cell surface into the cytosol. In this process, the plasma membrane undergoes massive local remodeling events as well as local changes in lipid composition mediated and regulated by the interplay of over 40 proteins that bind at specific time points at the membrane. One of these proteins is Epsin1. It is highly conserved across eukaryotic evolution and its structure reveals a tightly folded N-terminal ENTH (Epsin N-terminal homology) domain and an unstructured C-terminus. The ENTH domain has a compact globular structure mainly composed of helices. It is the membrane binding module and specifically binds to phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). The binding to PtdIns(4,5)P2 causes the formation of an amphipathic helix, which is associated with membrane deformation. In this thesis, the roles of additional phospholipids beside PtdIns(4,5)P2 in ENTH domain induced membrane deformation were investigated. Especially, phosphatidylserine (PtdSer) was identified to promote ENTH domain induced membrane tubulation by analysis of liposome morphology. Solid-state NMR studies revealed that the ENTH domain undergoes conformational changes in presence of PtdSer in the membrane. The generation of membrane tubules was accompanied by the formation of ENTH domain homo-oligomers. By mutating an arginine within the loop between helix 6 and helix 7 to an alanine (R114A) oligomerization as well as membrane deformation was inhibited. The ENTH domain was demonstrated to form homo-oligomers in presence of PtdSer by FRET studies and this particular loop was identified as the location for ENTH domain interprotein interaction site by MS/MS.
Keywords: ENTH domain; epsin; membrane deformation