title: Hydrophobic interaction chromatography on octyl sepharose--an approach for one-step platform purification of cyclodextrin glucanotransferases.
abstract: Cyclodextrin glucanotransferase (CGTase) from Bacillus circulans ATCC 21783 was concentrated by ultrafiltration and subsequently purified by hydrophobic interaction chromatography on Octyl Sepharose 4 fast flow. The matrix was able to bind selectively to the enzyme at a very low ammonium sulfate concentration of 0.67 M and enzyme desorption was performed by decreasing gradient of the salt. The overall recovery was 80% with 689-fold purity. CGTases derived from four soil isolates and Toruzyme, the commercial preparation of CGTase, also bound to Octyl Sepharose under similar conditions at 0.67 M and eluted at 0.55-0.5 M of ammonium sulfate. Octyl Sepharose chromatography can thus be used as a platform approach for purification of CGTases from various bacterial sources. Long stretches of sequence predominated by hydrophobic amino acids are reportedly present in the starch binding domains of CGTases. Starch binding experiments indicated the binding of the enzymes to the octyl matrix through these domains.
authors.affiliation: Department of Biotechnology, Manipal Life Sciences Centre, Manipal University, Manipal, Karnataka, India. premalatha_shetty@yahoo.co.in
authors.forename: Premalatha
authors.initials: P
authors.lastname: Shetty
authors.forename: Smitha
authors.initials: S
authors.lastname: Bhat
authors.forename: J L
authors.initials: JL
authors.lastname: Iyer
authors.forename: Srikant
authors.initials: S
authors.lastname: Shenoy
authors.forename: J S
authors.initials: JS
authors.lastname: Pai
authors.forename: K
authors.initials: K
authors.lastname: Satyamoorthy
mesh_terms: D001407:Bacillus / Q000201:enzymology*; D001665:Binding Sites; D002846:Chromatography, Affinity / Q000379:methods; D005964:Glucosyltransferases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D011485:Protein Binding; D012685:Sepharose / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: C027176:octyl-sepharose CL-4B; D012685:Sepharose; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase