expression and purification of functionally active recombinant human alpha 1-antitrypsin in methylotrophic yeast pichia pastoris
Authors
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.
similar resources
Expression and Purification of Functionally Active Recombinant Human Alpha 1-Antitrypsin in Methylotrophic Yeast Pichia pastoris
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 i...
full textElevating the expression level of biologically active recombinant human alpha 1-antitrypsin in Pichia pastoris
Background: Human alpha 1-antitrypsin (AAT) is a potent inhibitor of multiple serine proteases, and protects tissues against their harmful effects. Individuals with reduced or abnormal production of this inhibitor need intravenous administration of exogenous protein. In this study, we employed the methylotrophic (methanol utilizing) yeast Pichia pastoris (P. pastoris) as a preferential host for...
full textExpression of recombinant proteins in the methylotrophic yeast Pichia pastoris.
Protein expression in the microbial eukaryotic host Pichia pastoris offers the possibility to generate high amounts of recombinant protein in a fast and easy to use expression system. As a single-celled microorganism P. pastoris is easy to manipulate and grows rapidly on inexpensive media at high cell densities. Being a eukaryote, P. pastoris is able to perform many of the post-translational mo...
full textEvaluation of pH/buffering conditions effect on the optimization of Recombinant Human Erythropoietin expression in the methylotrophic yeast, Pichia pastoris
Expression of recombinant proteins and drugs in Pichia pastoris has been in development since the late 1980s and the number of recombinant proteins produced in P. pastoris has increased significantly in the past several years. Unlike bacteria, this strain is capable of producing complex proteins with post translational modifications such as correct folding, glycosylation, proteolytic maturation...
full textExpression, Purification and Characterization of Human Recombinant Galectin 3 in Pichia pastoris
Background: Over the past century, the areas of genomics, proteomics and lipids have captured the attention of investigators worldwide. Carbohydrates, have recently received increased attention through the expanding field of glycobiology; probably because they are very complex and not encoded in the genome. Objectives: The purpose of this study was to express and purify recombinant human galec...
full textHigh level expression, efficient purification, and bioactivity of recombinant human metallothionein 3 (rhMT3) from methylotrophic yeast Pichia pastoris.
Metallothionein 3 (MT3) is an important biochemical mediator regulating many physiological and pathophysiological processes including neuron cell protection, privation of reactive oxygen species-induced DNA damage, and protection against light induced retinal damage. In this study, a human gene encoding for MT3 with c-terminal extension of His6-tag was inserted into vector pPICZaA, and overexpr...
full textMy Resources
Save resource for easier access later
Journal title:
avicenna journal of medical biotechnologyجلد ۳، شماره ۳، صفحات ۱۲۷-۱۳۴
Hosted on Doprax cloud platform doprax.com
copyright © 2015-2023