STRUCTURE OF THE PLECTIN ACTIN BINDING DOMAIN
Institute of Molecular Biology, Slovak Academy of Sciences, Dúbravská cesta 21, 845 51 Bratislava 45, Slovak Republic
Plectin and its isoforms are versatile cytolinker proteins of very large size (molecular mass over 500 kDa) that are expressed in a wide variety of mammalian tissues and cell types. Biochemical data indicate that plectin plays an important role in cytoskeleton network organization and regulation, with consequences for viscoelastic properties of the cytoplasm and the mechanical integrity and resistance of cells and tissues. Defects in plectin genes cause autosomal recessive or dominant hereditary diseases, characterized by severe skin blistering with or without muscular dystrophy. Plectin has been well characterized biochemically and genetically. Electron microscopy revealed that the protein has a dumbbell-like structure comprising a central rod domain (approximately 2 000 Å long) flanked by N- and C- terminal globular domains. Each of these domains contains several subdomains to which binding sites for various interaction partners have been mapped. Actin-binding domain (ABD) of plectin is located in proximity to its N-terminus. It consists of two so called calponin homology (CH) 1 and 2 subdomains.
Crystals of the plectin ABD were grown by the hanging drop diffusion method. Modification of crystallization conditions resulted in two crystal forms. Data from crystal form I (P21) were collected at room temperature to 2.0 Å resolution and from crystal form II (P212121) at cryo temperature to 2.2 Å resolution on the EMBL beamlines at the DORIS storage ring, DESY Hamburg. The structure was solved by molecular replacement method using utrophin ABD (PDB code 1QAG) as search model. Structures of both crystal forms were refined with the program REFMAC5. Recombinant molecule of the plectin ABD is an α protein consisting of 245 residues which form 11 helices. The structure is almost identical with the fimbrin ABD in spite of relatively low amino-acid sequence identity (23 %) and differs from those of utrophin and dystrophin mainly in orientation of CH1 and CH2 subdomains.