Biophysical Society Conference | Estes Park 2023

Membrane Budding and Fusion

Wednesday Speaker Abstracts

CRYOEM STRUCTURES OF DYNAMIN PROTEINS INVOLVED IN MEMBRANE FISSION AND FUSION

Jenny E. Hinshaw 1 ; John R Jimah 2 ; Sarah B Nyenhuis 1 ; 1 NIH, NIDDK, Bethesda, MD, USA 2 Princeton University, Department of Molecular Biology, Princeton, NJ, USA

Dynamin superfamily proteins (DSPs) are present in all organisms and are broadly implicated in membrane remodeling, actin dynamics and innate immunity. DSPs are mechanochemical GTPases whose function is dependent on oligomerization of the protein and conformational changes that occur during the GTP hydrolysis cycle. Dynamin, the founding member, is crucial for endocytosis, synaptic membrane recycling, membrane trafficking within the cell, and cytokinesis. Based on our structural analysis of dynamin and studies from numerous other research groups, the emerging model entails dynamin assembling around the necks of budding vesicles as a helical polymer and upon GTP hydrolysis, undergoes a significant constriction that ultimately leads to membrane fission. Recently, we have determined high-resolution structures of assembled membrane-bound dynamin in two nucleotide states, GTP-bound and post-hydrolysis GDP bound, by cryoEM methods. Our data reveals conformational changes that allow for the constriction of dynamin tubes from >50 nm to 36 nm with an inner lumen of only 3.4 nm. In addition, we have solved the helical structure Opa1, a DSP involved in mitochondria fusion, in the presence and absence of nucleotide. These helical assemblies exhibit nucleotide-dependent dimerization of the GTPase domains, a hallmark of DSPs. In contrast to other DSPs, OPA1 contains several unique secondary structures in the paddle domain that strengthen its membrane association through monotopic membrane-inserting helices. Novel structural features of OPA1 shed light on the effects of pathogenic point mutations associated with Dominant Optic Atrophy. Further, mutations chosen to disrupt OPA1 assembly interfaces and membrane binding cause mitochondrial fragmentation in cell-based assays, demonstrating the biological relevance of these interactions.

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