title: Actin isoform-specific conformational differences observed with hydrogen/deuterium exchange and mass spectrometry.
abstract: Actin can exist in multiple conformations necessary for normal function. Actin isoforms, although highly conserved in sequence, exhibit different biochemical properties and cellular roles. We used amide proton hydrogen/deuterium (HD) exchange detected by mass spectrometry to analyze conformational differences between Saccharomyces cerevisiae and muscle actins in the G and F forms to gain insight into these differences. We also utilized HD exchange to study interdomain and allosteric communication in yeast-muscle hybrid actins to better understand the conformational dynamics of actin. Areas showing differences in HD exchange between G- and F-actins are areas of intermonomer contacts, consistent with the current filament models. Our results showed greater exchange for yeast G-actin compared with muscle actin in the barbed end pivot region and areas in subdomains 1 and 2 and for F-actin in monomer-monomer contact areas. These results suggest greater flexibility of the yeast actin monomer and filament compared with muscle actin. For hybrid G-actins, the muscle-like and yeastlike parts of the molecule generally showed exchange characteristics resembling their parent actins. A few exceptions were a peptide on top of subdomain 2 and the pivot region between subdomains 1 and 3 with muscle actin-like exchange characteristics although the areas were yeastlike. These results demonstrate that there is cross-talk between subdomains 1 and 2 and the large and small domains. Hybrid F-actin data showing greater exchange compared with both yeast and muscle actins are consistent with mismatched yeast-muscle interfaces resulting in decreased stability of the hybrid filament contacts.
authors: Department of Biochemistry, University of Iowa Carver College of Medicine, Iowa City, Iowa 52242, USA.
authors: Ema
authors: E
authors: Stokasimov
authors: Peter A
authors: PA
authors: Rubenstein
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000737:chemistry*; D000818:Animals; D003599:Cytoskeleton / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000737:chemistry; D003903:Deuterium / Q000737:chemistry; D006859:Hydrogen / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods; D008968:Molecular Conformation; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
chemical_list: D000199:Actins; D010455:Peptides; D020033:Protein Isoforms; D006859:Hydrogen; D003903:Deuterium; D003850:Deoxyribonuclease I