학술논문
Expression and Purification of Functionally Active Recombinant Human Alpha 1-Antitrypsin in Methylotrophic Yeast Pichia pastoris.
Document Type
Article
Source
Subject
*GENETIC engineering
*ALPHA 1-antitrypsin
*GENE expression
*RECOMBINANT proteins
*RESEARCH funding
*PROTEASE inhibitors
*METHODOLOGY
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Language
ISSN
2008-2835
Abstract
Human alpha 1-antitrypsin (AAT) cDNA was obtained from HepG2 cell lines. After PCR and construction of expression vector pPICZα-AAT, human AAT was expressed in the yeast Pichia pastoris (P.pastoris) in a secretary manner and under the control of inducible alcohol oxidase 1 (AOX1) promoter. The amount of AAT protein in medium was measured as 60 mg/l 72 hr after induction with methanol. Results indicated the presence of protease inhibitory function of the protein against elastase. Purification was done using His-tag affinity chromatography. Due to the different patterns of glycosylation in yeast and human, the recombinant AAT showed different SDS-PAGE patterns compared to that of serum-derived AAT while pI shifted from 4.9 in native AAT compared to 5.2 in recombinant AAT constructed in this study. [ABSTRACT FROM AUTHOR]