Biological activities of purified prolidase from pathogenic E.coli

Abstract

Prolidase is a ubiquitous enzyme that plays a chief role in the metabolism of proline-rich proteins. The goal of this study extraction and purify prolidase from E. coli and evaluate some applications such as anti-biofilm and anticancer. Hundred stool Samples were collected from infants with breastfeeding, non-vomiting, and non-diarrhea to isolate E.coli bacteria. A 16S rRNA gene (585 bp) was found in all isolates of E. coli via PCR identification. Depending on the qualitative method on prolidase agar, only 40 (80%) isolates could produce prolidase from 50 isolates that were considered non-pathogens, then only 32 isolates revealed different levels in prolidase production with specific activity equal to (2.1U/mg) of E.coli. MS12. Sucrose, casein, and 40Co were the chosen isolate's best conditions for producing prolidase. Cold acetone precipitation and dialysis were used to extract the enzyme, and DEAE-cellulose and the Sephadex G-150 column were used in purification with specific activity (2 U/ml) and (6.6 U/mg) protein. Prolidase showed the highest effect on biofilm at 500 μg/ml concentration against P. aeruginosa, then E. coli, 65% and 60.3% respectively. Brain Tumor Cell Line (A127), Colorectal Adenocarcinoma cells (CaCo-2), and Normal embryonic liver cell line (WRL-68) were used to test the prolidase effect on these cell lines. An assay of MTT was used to detect the inhibiter concentration (IC50) values and cytotoxic effect of purified prolidase. Keywords: E.coli, Prolidase, purification, antibiofilm, anticancer activity