title: Purification, crystallization and preliminary X-ray crystallographic analysis of squid heavy meromyosin.
abstract: All muscle-based movement is dependent upon carefully choreographed interactions between the two major muscle components, myosin and actin. Regulation of vertebrate smooth and molluscan muscle contraction is myosin based (both are in the myosin II class), and requires the double-headed form of myosin. Removal of Ca2+ from these muscles promotes a relatively compact conformation of the myosin dimer, which inhibits its interaction with actin. Although atomic structures of single myosin heads are available, the structure of any double-headed portion of myosin, including the ∼375 kDa heavy meromyosin (HMM), has only been visualized at low (∼20 Å) resolution by electron microscopy. Here, the growth of three-dimensional crystals of HMM with near-atomic resolution (up to ∼5 Å) and their X-ray diffraction are reported for the first time. These crystals were grown in off-state conditions, that is in the absence of Ca2+ and the presence of nucleotide analogs, using HMM from the funnel retractor muscle of squid. In addition to the crystallization conditions, the techniques used to isolate and purify this HMM are also described. Efforts at phasing and improving the resolution of the data in order to determine the structure are ongoing.
authors.affiliation: Rosenstiel Basic Medical Sciences Research Center, Brandeis University, 415 South Street, Waltham, MA 02454-9110, USA.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: O'Neall-Hennessey
authors.forename: Ludmila
authors.initials: L
authors.lastname: Reshetnikova
authors.forename: V S
authors.initials: VS
authors.lastname: Senthil Kumar
authors.forename: Howard
authors.initials: H
authors.lastname: Robinson
authors.forename: Andrew G
authors.initials: AG
authors.lastname: Szent-Györgyi
authors.forename: Carolyn
authors.initials: C
authors.lastname: Cohen
mesh_terms: D000818:Animals; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D003460:Crystallization; D018360:Crystallography, X-Ray; D049832:Decapodiformes / Q000737:chemistry*; D009132:Muscles / Q000737:chemistry*; D015879:Myosin Subfragments / Q000737:chemistry* / Q000302:isolation & purification; D055503:Protein Multimerization
chemical_list: D015879:Myosin Subfragments; D002118:Calcium
keywords: Loligo pealei; squid heavy meromyosin