title: Carrier free co-immobilization of alpha amylase, glucoamylase and pullulanase as combined cross-linked enzyme aggregates (combi-CLEAs): a tri-enzyme biocatalyst with one pot starch hydrolytic activity.
abstract: A tri-enzyme biocatalyst "combi-CLEAs" with starch hydrolytic activity was prepared from commercially available alpha amylase, glucoamylase and pullulanase preparations by aggregating enzymes with ammonium sulphate followed by cross-linking formed aggregates for 4.5h with 40 mM glutaraldehyde. The effects of precipitant type and cross-linking were studied and the biocatalyst was characterized. Scanning electron microscopy analysis showed that tri-enzyme biocatalyst was of spherical structure. For one pot starch hydrolytic activity, shift in optimum pH from 6 to 7 and temperature from 65 to 75 °C were observed after co-immobilization of enzymes. After one pot starch hydrolysis reaction in batch mode, 100%, 60% and 40% conversions were obtained with combi-CLEAs, separate CLEAs mixture and free enzyme mixture, respectively. Co-immobilization also enhanced the thermal stability of enzymes. Finally, the catalytic activity of enzymes in combi-CLEAs during one pot starch hydrolysis was well maintained up to five cycles without performance changes.
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India. Electronic address: sachintalekar7@gmail.com.
authors.forename: Sachin
authors.initials: S
authors.lastname: Talekar
authors.affiliation: Chemical Engineering and Process Development Division, CSIR-National Chemical Laboratory, Pune 411 008, India.
authors.forename: Amol
authors.initials: A
authors.lastname: Pandharbale
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India.
authors.forename: Mayur
authors.initials: M
authors.lastname: Ladole
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India.
authors.forename: Shamraja
authors.initials: S
authors.lastname: Nadar
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India.
authors.forename: Mosin
authors.initials: M
authors.lastname: Mulla
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India.
authors.forename: Kshitija
authors.initials: K
authors.lastname: Japhalekar
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India.
authors.forename: Kishori
authors.initials: K
authors.lastname: Pattankude
authors.affiliation: Department of Biotechnology Engineering, Kolhapur Institute of Technology's College of Engineering, Kolhapur 416 234, India.
authors.forename: Devika
authors.initials: D
authors.lastname: Arage
mesh_terms: D055162:Biocatalysis; D004795:Enzyme Stability; D005087:Glucan 1,4-alpha-Glucosidase / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006868:Hydrolysis; D013696:Temperature; D000516:alpha-Amylases / Q000378:metabolism*
chemical_list: D006026:Glycoside Hydrolases; D000516:alpha-Amylases; D005087:Glucan 1,4-alpha-Glucosidase; C100181:pullulanase
keywords: Alpha amylase; Combined cross-linked enzyme aggregates (combi-CLEAs); Glucoamylase; One pot starch hydrolysis; Pullulanase