Heterologous Expression, Purification and
Determination of Activity of new Carboxypepetidase
 from Cohnella Ao1

Seyed Mahdi Naeemi¹, Saeed Aminzadeh^2*, Soyar Sari¹,
Fahimeh Nemati³, Maryam naseroleslami¹
1- Department of Cellular and Molecular Biology, Faculty of Advanced Science and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran
2- Bioprocess Engineering Group, Institute of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
3- Department of Biotechnology, Faculty of Advanced Science and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran.

Aim and Background: Proteolytic enzymes are essential for the survival of living organisms by irreversible hydrolysis of peptide bond. Carboxypeptidases are proteolytic enzymes found in many organisms and are widely used in various industries, such as food and pharmaceuticals. Aims of this study are expression of carboxypeptidase of chohnella A01, purification and determination of its activity.

Materials and methods: The gene of carboxypeptidase of cohnella A01 has been cloned in pET26b(+) vector. Competent cells were prepared from E. coli BL21, plasmids were extracted and transformed to expression host. E. coli BL21 was cultured in LB medium and protein expression was induced by IPTG. After that cell harvested and recombinant protein was purified by Ni-TNA resin. 

Results: The expressed recombinant enzyme is a glutamate carboxypeptidase that capable of separating glutamate from folic acid.

Conclusion: Hence carboxypeptidase G has medicinal application and recombinant carboxypeptidase from cohnella A01 is a thermostable enzyme, after optimization, can be use it for medicinal application.

Keywords: Carboxypeptidase, Cohnella, Folic Acid, Glutamate, IAU science