title: Functional expression of trypsin from Streptomyces griseus by Pichia pastoris.
abstract: In the present study, the genes encoding trypsinogen and active trypsin from Streptomyces griseus were both cloned and expressed in the methylotrophic yeast Pichia pastoris with the α-factor secretion signal under the control of the alcohol oxidase promoter. The mature trypsin was successfully accumulated extracellularly in soluble form with a maximum amidase activity of 6.6 U ml(-1) (batch cultivation with flask cultivation) or 14.4 U ml(-1) (fed-batch cultivation with a 3-l fermentor). In contrast, the recombinant trypsinogen formed inclusion bodies and no activity was detected. Replacement of the trypsin propeptide Ala-Pro-Asn-Pro confirmed that its physiological function was as a repressor of activity. More importantly, our results proved that the propeptide inhibited the activity of trypsinogen after its successful folding.
authors: The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China.
authors: Zhenmin
authors: Z
authors: Ling
authors: Tengbo
authors: T
authors: Ma
authors: Jianghua
authors: J
authors: Li
authors: Guocheng
authors: G
authors: Du
authors: Zhen
authors: Z
authors: Kang
authors: Jian
authors: J
authors: Chen
mesh_terms: D000581:Amidohydrolases / Q000378:metabolism; D019149:Bioreactors; D004950:Esterases / Q000378:metabolism; D005285:Fermentation; D002479:Inclusion Bodies / Q000378:metabolism; D010843:Pichia / Q000235:genetics* / Q000378:metabolism*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015203:Reproducibility of Results; D013305:Streptomyces griseus / Q000235:genetics / Q000378:metabolism*; D014357:Trypsin / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014362:Trypsinogen / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D014362:Trypsinogen; D004950:Esterases; D014357:Trypsin; D000581:Amidohydrolases