title: Purification and characterization of recombinant Streptomyces griseus aminopeptidase.
abstract: Recombinant Streptomyces griseus aminopeptidase (SGAP) was produced using Cangene's expression system, CANGENUS. This heat-stable aminopeptidase with an N-terminal Ala-Pro-Asp-Ile-Pro-Leu-Ala-Asn-Val-Lys-Ala sequence was purified from 16L of Streptomyces lividans fermentation supernatant with high purity and 19.5% recovery rate. This was achieved by the combination of hydrophobic-interaction and size-exclusion chromatographic procedures. The calcium-activated zinc metalloprotein demonstrated no loss of activity at -20 degrees C for at least 8 weeks in both liquid and freeze-dried formulations. The recombinant SGAP showed an apparent molecular mass of 31 kDa by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and 26.8 kDa by gel filtration. The simple, high-yield, inexpensive purification method with few intermediate steps provides a novel and practical procedure for large-scale production of active recombinant S. griseus aminopeptidase.
authors.affiliation: Department of Biotechnology Research, Cangene Corporation, Mississauga, ON, Canada.
authors.forename: Sally Xueying
authors.initials: SX
authors.lastname: Ni
authors.forename: Douglas
authors.initials: D
authors.lastname: Cossar
authors.forename: Adrian
authors.initials: A
authors.lastname: Man
authors.forename: Karen
authors.initials: K
authors.lastname: Norek
authors.forename: David
authors.initials: D
authors.lastname: Miller
authors.forename: Craig
authors.initials: C
authors.lastname: Kearse
authors.forename: Vadim
authors.initials: V
authors.lastname: Tsvetnitsky
mesh_terms: D000626:Aminopeptidases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D005285:Fermentation; D005822:Genetic Vectors; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013305:Streptomyces griseus / Q000201:enzymology* / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D000626:Aminopeptidases