title: Expression in Escherichia coli of human lamins A and C: influence of head and tail domains on assembly properties and paracrystal formation.
abstract: We have expressed in Escherichia coli cDNA corresponding to human lamins A and C, together with a number of fragments produced using site-specific mutagenesis. The proteins produced in this way were characterised both biochemically and ultrastructurally, and appeared to retain their native conformation. Crosslinking showed that all fragments formed 4-chain molecular dimers ('tetramers') analogous to those formed by intact intermediate filament proteins. Shadowed preparations showed the presence of rod-like particles that closely resembled those observed for other intermediate filament proteins and their proteolytically prepared rod domains. Moreover, the expressed lamins and a series of fragments in which different domains had been deleted formed paracrystals similar to those observed with native material. Deletion of either the N- or C-terminal non-helical domains altered the solubility and aggregation properties of the expressed protein, indicating that both domains have a role in lamin assembly.
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge, UK.
authors.forename: R D
authors.initials: RD
authors.lastname: Moir
authors.forename: A D
authors.initials: AD
authors.lastname: Donaldson
authors.forename: M
authors.initials: M
authors.lastname: Stewart
mesh_terms: D003001:Cloning, Molecular; D003460:Crystallization; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D034882:Lamins; D008854:Microscopy, Electron; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D012995:Solubility
chemical_list: D034882:Lamins; D009687:Nuclear Proteins