title: Structural evidence for non-canonical binding of Ca2+ to a canonical EF-hand of a conventional myosin.
abstract: We have previously identified a single inhibitory Ca2+-binding site in the first EF-hand of the essential light chain of Physarum conventional myosin (Farkas, L., Malnasi-Csizmadia, A., Nakamura, A., Kohama, K., and Nyitray, L. (2003) J. Biol. Chem. 278, 27399-27405). As a general rule, conformation of the EF-hand-containing domains in the calmodulin family is "closed" in the absence and "open" in the presence of bound cations; a notable exception is the unusual Ca2+-bound closed domain in the essential light chain of the Ca2+-activated scallop muscle myosin. Here we have reported the 1.8 A resolution structure of the regulatory domain (RD) of Physarum myosin II in which Ca2+ is bound to a canonical EF-hand that is also in a closed state. The 12th position of the EF-hand loop, which normally provides a bidentate ligand for Ca2+ in the open state, is too far in the structure to participate in coordination of the ion. The structure includes a second Ca2+ that only mediates crystal contacts. To reveal the mechanism behind the regulatory effect of Ca2+, we compared conformational flexibilities of the liganded and unliganded RD. Our working hypothesis, i.e. the modulatory effect of Ca2+ on conformational flexibility of RD, is in line with the observed suppression of hydrogen-deuterium exchange rate in the Ca2+-bound form, as well as with results of molecular dynamics calculations. Based on this evidence, we concluded that Ca2+-induced change in structural dynamics of RD is a major factor in Ca2+-mediated regulation of Physarum myosin II activity.
authors.affiliation: Department of Biochemistry, Eötvös Loránd University, Budapest H-1117, Hungary.
authors.forename: Judit E
authors.initials: JE
authors.lastname: Debreczeni
authors.forename: László
authors.initials: L
authors.lastname: Farkas
authors.forename: Veronika
authors.initials: V
authors.lastname: Harmat
authors.forename: Csaba
authors.initials: C
authors.lastname: Hetényi
authors.forename: István
authors.initials: I
authors.lastname: Hajdú
authors.forename: Péter
authors.initials: P
authors.lastname: Závodszky
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kohama
authors.forename: László
authors.initials: L
authors.lastname: Nyitray
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry; D002412:Cations; D018360:Crystallography, X-Ray; D003903:Deuterium / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D006859:Hydrogen / Q000737:chemistry; D007700:Kinetics; D008024:Ligands; D008958:Models, Molecular; D008968:Molecular Conformation; D018482:Muscle, Skeletal / Q000378:metabolism; D024462:Myosin Type II / Q000737:chemistry; D009218:Myosins / Q000737:chemistry*; D010804:Physarum / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D055672:Static Electricity; D013997:Time Factors
chemical_list: D002147:Calmodulin; D002412:Cations; D008024:Ligands; D006859:Hydrogen; D003903:Deuterium; D024462:Myosin Type II; D009218:Myosins; D002118:Calcium