IMPROVEMENT OF DIFFRACTION QUALITY UPON REHYDRATION OF DEHYDRATED ICOSAHEDRAL ENTEROCOCCUS-FAECALIS PYRUVATE-DEHYDROGENASE CORE CRYSTALS

Members of the family of 2-oxoacid dehydrogenase multienzyme complexes catalyze the oxidative decarboxylation of alpha-keto acids and are am ong the most remarkable enzymatic ma chineries in the living cell. The se multienzyme complexes combine a highly symmetric (cubic or icosahed ral) core with a dynamic and flexible arrangement of numerous subunits and domains surrounding the core. The center of the complex is formed by either 24 or 60 copies of dihydrolipoamide acetyltransferase (E2)- a multidomain enzyme. The hollow icosahedral cores are composed of 60 identical subunits of the catalytic domain of E2 with a molecular weig ht of about 1.8 million Da. Bipyramidal crystals suitable for X-ray di ffraction of the icosahedral core of the pyruvate dehydrogenase multie nzyme complex from Enterococcus faecalis were grown up to 0.7 mm in ea ch dimension. The crystals belong to space group R32 with a = b = 244. 3 Angstrom and c = 920.9 Angstrom (hexagonal setting), and have a solv ent content of 73%. The asymmetric unit contains one-third of the mole cule, i.e., 20 of the 60 subunits. Initial X-ray crystallographic data to 7 Angstrom resolution were collected at cryotemperatures at synchr otron facilities. Interestingly, the diffraction was improved signific antly upon rehydrating dehydrated crystals and extended to 4.2 Angstro m.