High-level extracellular secretion of organophosphorous hydrolase of Flavobacterium sp. in Escherichia coli BL21(DE3)pLysS

• Published in: Biotechnology and applied biochemistry 2016-11, Vol.63 (6), p.870-876
• Main Authors: Falahati-Pour, Soudeh Khanamani, Lotfi, Abbas Sahebghadam, Ahmadian, Gholamreza, Baghizadeh, Amin, Behroozi, Reza, Haghighi, Fereshteh
• Format: Article
• Language: English
• Subjects: Aryldialkylphosphatase - chemistry; Aryldialkylphosphatase - genetics; Aryldialkylphosphatase - secretion; Biochemistry; Biotechnology; Cleavage; Cobalt - pharmacology; Codon - genetics; Culture; Culture Media - chemistry; Dose-Response Relationship, Drug; Escherichia coli; Escherichia coli - cytology; Escherichia coli - drug effects; Escherichia coli - genetics; Escherichia coli - growth & development; Escherichia coli BL21(DE3)pLysS; Extracellular Space - metabolism; Flavobacterium; Flavobacterium - enzymology; Flavobacterium - genetics; Genetic Engineering - methods; Glycine - pharmacology; Isopropyl Thiogalactoside - pharmacology; organophosphorus hydrolase; Peptides; Plasmids - genetics; Protein Sorting Signals; Recombinant; secretion; Secretions; Tat signal peptide; Time Factors
• ISSN: 0885-4513; 1470-8744
• DOI: 10.1002/bab.1438

Organophosphorus (OPs) compounds are widely used in many pesticides, insecticides, and chemical nerve agents. These compounds are hazardous for humans and the environment. There are many reports on detoxification of these compounds, among them enzymatic cleavage of these compounds with organophosphorus hydrolase (OPH) has been taken into more consideration. Several studies have been performed to improve OPH secretion in Escherichia coli by different signal peptides, but have not been successful. In this study, to achieve the extracellular secretion of OPH in E. coli, the complete opd gene along with its native signal peptide was codon optimized and expressed in E. coli BL21(DE3)pLysS. The culture medium showed OPH activity after 2, 4, and 6 H of induction time. The extracellular secretion of OPH was also confirmed by SDS‐PAGE and Western blot analysis. The effects of different factors in growth medium were also investigated regarding expression and extracellular secretion of OPH. It appears that the secretion of OPH into the extracellular medium is highly affected by culture conditions. Therefore, our results revealed that the recombinant OPH was successfully secreted into the extracellular medium. This secretion system can be considered as a high efficiency biocatalyst for detoxification of OPs compounds.