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Cloning, Expression, And Activity Analysis Of Human Cathepsin C In The Yeast Pichia Pastoris, Cenk Dağlioğlu
Cloning, Expression, And Activity Analysis Of Human Cathepsin C In The Yeast Pichia Pastoris, Cenk Dağlioğlu
Turkish Journal of Biology
The yeast Pichia pastorisexpression system was investigated for the production of human cathepsin C (CatC) recombinant protein. The full-length CatC cDNA, corresponding to amino acids 12-475, was synthesized from interleukin-2 (IL-2) stimulated human peripheral blood mononuclear cells and subcloned in the pGEM-T cloning vector. After confirming the DNA sequence of the insert, the gene was cloned into the pPICZ\alphaA expression vector under the control of the methanol-inducible alcohol oxidase (AOX1) promoter and transformed to P. pastoris X-33 cells. The expressed protein was secreted into the culture medium through the alpha-factor mating signal sequence of the expression vector. Analysis of the …
Cloning, Expression, And Characterization Of Human Brain Acetylcholinesterase In Escherichia Coli Using A Sumo Fusion Tag, Hami̇d Ceylan, Orhan Erdoğan
Cloning, Expression, And Characterization Of Human Brain Acetylcholinesterase In Escherichia Coli Using A Sumo Fusion Tag, Hami̇d Ceylan, Orhan Erdoğan
Turkish Journal of Biology
The molecular structure of acetylcholinesterase (AChE) attracts interest because of its versatility and significant role in the cholinergic system. The main purpose of the present study was to clone a full-length cDNA sequence of human brain acetylcholinesterase (hAChE) into pET SUMO vector and express it successfully. The integrity of the constructed plasmid was confirmed by cross PCR. This recombinant construct was expressed in Escherichia coli BL21 (DE-3). In this work, we produced hexahistidine (6xHis) tagged fusion protein by isopropyl β-D-1-thiogalactopyranoside (IPTG) induction and purified using nickel (Ni2+) affinity chromatography. Using anti-His antibody, we detected ~90 kDa fusion protein. The expression …