EXTRACTION AND PURIFICATION OF TWO OUTER MEMBRANE PROTEINS (PORINS) FROM KLEBSIELLA PNEUMONIAE LOCAL ISOLATE

Abstract

Background: The porins are present in large amounts in the outer membrane of gram negative bacteria and form water-filled channels that permit the diffusion of small hydrophilic solutes across the outer membrane. Porins are generally divided into two classes: nonspecific porins (e.g., OmpC and OmpF), which permit the general diffusion of small polar molecules (600 Da), and specific porins (e.g., LamB), which facilitate the diffusion of specific substrates.Objective: To purify and characterize outer membrane proteins (porins) from a local isolate of Klebsiella pneumoniae. Materials and methods: An identified local isolate of Klebsiella pneumoniae was used as a primary source for the isolation and purification of porins. Outer membrane protein (porins) was purified and characterized and the contaminating lipopolysaccharides (LPS) were detected by thiobarbituric acid assay. Results: The final preparation contained porins in a concentration of 3.2 mg/ml. The results of electrophoretic separation revealed that porins appeared as two distinct bands with molecular weights of porins were estimated to be 35 and 36 kDa, respectively. Conclusions: Porins were expressed by the local isolate of Klebsiella pneumoniae with molecular weights highly similar to that of porins preparations produced by other gram negative bacteria and Klebsiella pneumoniae expressed two types of porins under standard laboratory conditions.Keywords: Porins, Thiobarbituric acid, Gel filtration chromatography, Ketodeoxyoctinate.