Biophysical Society Thematic Meeting| Padova 2019

Quantitative Aspects of Membrane Fusion and Fission

Poster Abstracts

23-POS Board 23 BIOCHEMICAL AND STRUCTURAL ANALYSIS OF A HUMAN DYNAMIN 2 MUTANT, R719W, ASSOCIATED WITH HEREDITARY SPASTIC PARAPLEGIA (HSP) John Jimah 1 ; Abigail Stanton 1 ; Jenny Hinshaw 1 ; Lieza Chan 1 ; 1 National Institutes of Health, Structural Cell Biology Section, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland, USA Classical dynamin GTPases (dynamin 1, 2 and 3) mediate membrane fission during endocytosis. Currently, all known disease-associated mutations in dynamins occur in dynamin 2. One such mutation, R719W, is linked to HSP, a disease that causes muscle weakness and stiffness resulting in the inability to walk. The objective of this study is to uncover the molecular mechanism of dynamin 2 in normal and disease states. Structural modeling of dynamin 2 situates residue R719 in the bundle signaling element (BSE), a three-helix bundle that links dynamin’s GTPase domain to the stalk domain. Upon GTP hydrolysis by wt dynamin, the BSE swings downward by 68 o relative to the GTPase domain (powerstroke) resulting in rearrangements in the polymer that leads to constriction of dynamin-lipid tubules from an inner luminal diameter of 7 nm to 4 nm. Cells expressing the R719W mutant are deficient in endocytosis, which may be due to reduced GTP hydrolysis-dependent powerstroke. Our experimental methods involve obtaining structures of dynamin 2 wt and R719W mutant assembled on lipid membranes to elucidate the mechanism of assembly and membrane constriction. Preliminary results from cryo-EM studies reveal that the extent of membrane constriction may differ between dynamin 2 wt and R719W. This work will increase our understanding of the structural features of dynamins necessary for membrane fission and the molecular basis of dynamin-associated diseases.

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