Cloning, expression, purification, crystallization and preliminary X-ray diffraction studies of the catalytic domain of a hyperthermostable endo-1,4-β-D-mannanase fromThermotoga petrophilaRKU-1

Abstract

Endo-1,4-β-D-mannanases play key roles in seed germination and fruit ripening and have recently received much attention owing to their potential applications in the food, detergent and kraft pulp industries. In order to delineate their structural determinants for specificity and stability, X-ray crystallographic investigations combined with detailed functional studies are being performed. In this work, crystals of the catalytic domain of a hyperthermostable endo-1,4-β-D-mannanase fromThermotoga petrophilaRKU-1 were obtained from three different conditions, resulting in two crystalline forms. Crystals from conditions with phosphate or citrate salts as precipitant (CryP) belonged to space groupP212121, with unit-cell parametersa= 58.76,b= 87.99,c= 97.34 Å, while a crystal from a condition with ethanol as precipitant (CryE) belonged to space groupI212121, with unit-cell parametersa= 91.03,b= 89.97,c= 97.89 Å. CryP and CryE diffracted to resolutions of 1.40 and 1.45 Å, respectively.