Cloning, expression, and characterization of human brain acetylcholinesterase in Escherichia coli using a SUMO fusion tag

Hamid CEYLAN; Orhan ERDOĞAN

Makale detayları

Cloning, expression, and characterization of human brain acetylcholinesterase in Escherichia coli using a SUMO fusion tag

Yazarlar:

Hamid CEYLAN | Orhan ERDOĞAN

Yayıncı:

TUBITAK

Ana Başlıklar:

Yaşam bilimleri

Dergi:

Turkish Journal of Biology 2017 Vol.41 Issue 1, pp.77-87

Belge Tipi:

Makale

Yayın Tarihi:

1/2017

Belge Dili:

İngilizce

Document ID (Org No.):

2000811468

Sıklık:

İki Aylık

eISSN:

1303-6092

pISSN:

1300-0152

Özet:

 

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 of the hAChE gene in a microbial host resulted in good biological activity. Using the Ellman method, the recombinant AChE exhibited activity with optima at pH 9.0 glycine-NaOH buffer and room temperature. Kinetic parameters, KM and Vmax, were determined as 0.63 and 0.69, respectively.

Makale detayları

Cloning, expression, and characterization of human brain acetylcholinesterase in Escherichia coli using a SUMO fusion tag

Yazarlar: Hamid CEYLAN | Orhan ERDOĞAN

Yayıncı: TUBITAK

Ana Başlıklar: Yaşam bilimleri

Dergi: Turkish Journal of Biology 2017 Vol.41 Issue 1, pp.77-87

Belge Tipi: Makale

Yayın Tarihi: 1/2017

Belge Dili: İngilizce

Document ID (Org No.): 2000811468

Sıklık: İki Aylık

eISSN: 1303-6092

pISSN: 1300-0152

Sayı

Yazarlar:

Hamid CEYLAN | Orhan ERDOĞAN

Yayıncı:

TUBITAK

Ana Başlıklar:

Yaşam bilimleri

Dergi:

Turkish Journal of Biology 2017 Vol.41 Issue 1, pp.77-87

Belge Tipi:

Makale

Yayın Tarihi:

1/2017

Belge Dili:

İngilizce

Document ID (Org No.):

2000811468

Sıklık:

İki Aylık

eISSN:

1303-6092

pISSN:

1300-0152