Purification Methods
This trehalase is purified by solubilising in Triton X-100 and sodium deoxycholate, and submitting to gel filtration, ion-exchange chromatography, conA-Sepharose chromatography, phenyl-Sepharose CL-4B hydrophobic interaction chromatography, Tris-Sepharose 6B affinity and hydrolyapatite chromatography. Activity is increased 3000-fold. [Yoneyama Arch Biochem Biophys 255 168 1987.] T4-RNA ligase (from bacteriophage-infected E.coli ) Mr 43,500, [EC 6.5.1.3 for RNA lyase]. This ligase is purified by differential centrifugation and separation on a Sephadex A-25 column, then through hydroxylapatite and DEAE-glycerol using Aff-Gel Blue to remove DNAase activity. (Greater than 90% of the protein in the enzyme preparation migrated as a single band on gradient polyacrylamide gels containing SDS during electrophoresis.) [McCoy et al. Biochim Biophys Acta 562 149 1979.]
