Expression, Solubilization, Refolding and Final Purification of Recombinant Proteins as Expressed in the form of "Classical Inclusion Bodies" in E. coli.

Escherichia coli has been most widely used for production of the recombinant proteins. Over-expression of the recombinant proteins is the mainspring of the inclusion bodies formation. The refolding of these proteins into bioactive forms is cumbersome and partly time-consuming. In the present study, we reviewed and discussed most issues regarding the recovery of "classical inclusion bodies" by focusing on our previous experiences. Performing proper methods of expression, solubilization, refolding and final purification of these proteins, would make it possible to recover higher amounts of proteins into the native form with appropriate conformation. Generally, providing mild conditions and proper refolding buffers, would lead to recover more than 40% of inclusion bodies into bioactive and native conformation.

The distribution of pathogenic and toxigenic genes among MRSA and MSSA clinical isolates.

Staphylococcus aureus (S. aureus) is considered as a notorious nosocomial pathogen among hospitalized patients and community-dwelling subjects. Its increasing morbidity and mortality is believed to be due to antibiotic resistance. However, the data concerning molecular properties of infecting strains are few. In this study, a total of 192 S. aureus strains, including 88 (45.8%) meticillin-sensitive S. aureus (MSSA) and 104 (54.2%) meticillin-resistant S. aureus (MRSA) were recovered from clinical samples. The prevalence of subtypes containing staphylococcal cassette chromosome mec (SSCmec), staphylococcal enterotoxins (SEs), toxic shock syndrome toxin (TSST) and exfoliative toxin was assessed by PCR. Antibiotic susceptibility pattern and vancomycin resistance of each isolate were evaluated by disk diffusion method and micro-dilution method, respectively. 9 (2.3%) strains required MIC > 2 mg/l of vancomycin, which significantly increased among multi drug resistant (MDR), MRSA and SCCmec type III strains (p < 0.05). 171 (89%), 140 (72.91%), 7 (3.6), 78 (48.6%), 5 (2.6%), 151 (78.64%), 129 (67.18%), 178 (92.7%) and 15 (7.8%) of 192 isolates harbored mecA, entA, entB, entC, entD, entE, eta, etb and tsst-1 genes, respectively. 31 (16.14%), 5 (2.6%), 95 (49.48%) and 7 (3.64%) of 192 isolates carried SCCmec type I, II, III and IV, respectively. We found a significantly higher rate of MRSA and resistance to all tested antibiotics, except to penicillin G, kanamycin and linezolide among the SCCmec type III class (p < 0.05). According to our findings, MSSA isolates should be taken as seriously as MRSA strains due to the potential presence of broad spectrum virulence factor genes.