Crystallization and preliminary X-ray analysis of human transglutaminase 3from zymogen to active form

Transglutaminases (TGases; protein-glutamine-glutamyl-transferases) are a l arge family of calcium-dependent acyl-transfer enzymes that catalyze the fo rmation of covalent cross links in proteins. Of these, the "epidermal" or " hair follicle" TGase 3 isoform is critically involved in barrier formation in epithelia. It is a zymogen, requiring proteolytic activation to achieve maximal specific activity. In order to understand its structure and functio n, we have devised methods for the rapid large-scale expression of the TGas e 3 zymogen in the baculovirus system, and here we describe the purificatio n of the zymogen and activated forms. We describe methods for the formation of high-quality, well-diffracting crystals within 3-5 days, using both dio xane and beta -octylglucoside to overcome severe twinning problems. The cry stal of the zymogen belongs to the triclinic space group P1 and diffracts t o 2.2-Angstrom resolution, and the crystal of the active form belongs to th e P2(1) space group at 2.7-Angstrom resolution.