Purification, crystallization and preliminary X-ray analysis of the plectin actin-binding domain.
Plectin is an abundantly expressed cytoskeletal crosslinking protein of enormous size (>500 kDa) and multiple functions. It represents one of the many members of a large family of actin-binding proteins. The actin-binding domain of mouse plectin was expressed in Escherichia coli and purified to homogeneity. Crystals of the actin-binding domain of plectin were prepared by the hanging-drop method. They belong to space group P2(1), with unit-cell parameters a = 55.92, b = 108.92, c = 63.75 A, beta = 115.25 degrees. Data from a single crystal were collected to 2.0 A resolution at room temperature using synchrotron radiation at EMBL, Hamburg. The asymmetric unit contains two molecules of the protein, which corresponds to V(M) = 3.06 A(3) Da(-1) and a solvent content of 60%. The structure was solved by the molecular-replacement method. In addition, the preparation of selenomethionine-derivative crystals is described.
Urbániková
L'ubica
L
Institute of Molecular Biology, Slovak Academy of Sciences, Dúbravská cesta 21, 842 51 Bratislava, Slovak Republic. umbiurbi@savba.savba.sk
Janda
Lubomír
L
Popov
Alexander
A
Wiche
Gerhard
G
Sevcík
Jozef
J
D000199:Actins; D000818:Animals; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D007381:Intermediate Filament Proteins / Q000737:chemistry* / Q000302:isolation & purification; D051379:Mice; D015394:Molecular Structure; D051190:Plectin; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D012645:Selenomethionine
D000199:Actins; D007381:Intermediate Filament Proteins; C495333:Plec protein, mouse; D051190:Plectin; D011994:Recombinant Proteins; D012645:Selenomethionine