Skip Navigation
Skip to contents

PHRP : Osong Public Health and Research Perspectives

OPEN ACCESS
SEARCH
Search

Author index

Page Path
HOME > Articles and issues > Author index
Search
Rezvan Moniri 1 Article
Expression of Recombinant pET22b-LysK-Cysteine/Histidine-Dependent Amidohydrolase/Peptidase Bacteriophage Therapeutic Protein in Escherichia coli BL21 (DE3)
Hamed Haddad Kashani, Rezvan Moniri
Osong Public Health Res Perspect. 2015;6(4):256-260.   Published online August 31, 2015
DOI: https://doi.org/10.1016/j.phrp.2015.08.001
  • 1,796 View
  • 19 Download
  • 17 Citations
AbstractAbstract PDF
Objectives
Bacteriophage-encoded endolysins are a group of enzymes that act by digesting the peptidoglycan of bacterial cell walls. LysK has been reported to lyse live staphylococcal cultures. LysK proteins containing only the cysteine/histidine-dependent amidohydrolase/peptidase (CHAP) domain has the capability to show lytic activity against live clinical staphylococcal isolates, including methicillin-resistant Staphylococcus aureus (MRSA). The aim of this study was to clone and express LysK-CHAP domain in Escherichia coli BL21 (DE3) using pET22b as a secretion vector. The pET22b plasmid was used, which encoded a pelB secretion signal under the control of the strong bacteriophage T7 promoter.
Methods
The E. coli cloning strains DH5α and BL21 (DE3) were grown at 37°C with aeration in the Luria-Bertani medium. A plasmid encoding LysK-CHAP in a pET22b backbone was constructed. The pET22b vector containing LysK-CHAP sequences were digested with NcoI and HindIII restriction enzymes. Cloning accuracy was confirmed by electrophoresis. The pET22b-LysK plasmid was used to transform the E. coli strain BL21. Isopropyl β-d-1-thiogalactopyranoside (IPTG) was added to a final concentration of 1mM to induce T7 RNA polymerase-based expression. Finally, western blot confirmed the expression of target protein.
Results
In this study, after double digestion of pEX and pET22b vectors with HindIII and NcoI, LysK gene was cloned into two HindIII and NcoI sites in pET22b vector, and then transformed to E. coli DH5α. Cloning was confirmed with double digestion and analyzed with agarose gel. The recombinant pET22b-LysK plasmid was transformed to E. coli BL21 and the expression was induced by IPTG. The expression was confirmed by Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting method. Observation of a 28.5 kDa band confirmed LysK protein expression.
Conclusion
In the present study, LysK-CHAP domain was successfully cloned and expressed at the pET22b vector and E. coli BL21 (DE3).

Citations

Citations to this article as recorded by  
  • Chemotherapeutic Strategies for Combating Staphylococcus aureus Infections
    Namita Sharma, Anil Kumar Chhillar, Sweety Dahiya, Aruna Punia, Pooja Choudhary, Prity Gulia, Akanksha Behl, Mehak Dangi
    Mini-Reviews in Medicinal Chemistry.2022; 22(1): 26.     CrossRef
  • Antimicrobial peptides of the vaginal innate immunity and their role in the fight against sexually transmitted diseases
    H. Madanchi, M. Shoushtari, H.H. Kashani, S. Sardari
    New Microbes and New Infections.2020; 34: 100627.     CrossRef
  • Assessment of Relationship Between Expression of Survivin Protein and Histopathology Diagnosis and Malignancy Severity in Colon Specimen
    Amin Jourabchin, Tahereh Mazoochi, Hamed Haddad Kashani, Tahereh Khamechian
    Journal of Gastrointestinal Cancer.2020; 51(1): 76.     CrossRef
  • The effect of calcium on the adhesion of Streptococcus mutans to Human Gingival Epithelial Cells in the presence of probiotic bacteria Lactobacillus plantarum and Lactobacillus salivarius
    Mahboobeh Mehrabani Natanzi, Fatemeh Soleimanifard, Hamed Haddad Kashani, Mohammad Javad Azadchehr, Ahmadreza Mirzaei, Zohre Khodaii
    Gene Reports.2020; 20: 100710.     CrossRef
  • Expression and characterization of family 40 Carbohydrate Binding Module (CBM) from Vibrio cholerae Non-O1 sialidase
    Gogula Selvi Asang, Shadariah Mamat, Nadiawati Alias, Asmad Kari
    Asia Pacific Journal of Molecular Biology and Biot.2020; : 26.     CrossRef
  • Effect of intraoperative dexmedetomidine infusion during functional endoscopic sinus surgery: a prospective cohort study
    Mohammad Reza Fazel, Zeynab Sadat Ahmadi, Hossein Akbari, Fahimeh Abam
    Patient Safety in Surgery.2020;[Epub]     CrossRef
  • Comparison of a novel herbal skin care ointment with regular ointments to treat skin around the abdominal stoma: A clinical trial study
    Maryam Hajikari, Soheila Mojdeh, Mohsen Shariari
    Polish Annals of Medicine.2019;[Epub]     CrossRef
  • Evaluation of the serum sex hormones levels and alkaline phosphatase activity in rats’ testis after administering of berberine in experimental varicocele
    Hamed Najaran, Hassan Hassani Bafrani, Hamid Rashtbari, Fatemeh Izadpanah, Mohammad Reza Rajabi, Hamed Haddad Kashani, Abouzar Mohammadi
    Oriental Pharmacy and Experimental Medicine.2019; 19(2): 157.     CrossRef
  • Effect of melatonin in reducing second-generation antipsychotic metabolic effects: A double blind controlled clinical trial
    Mansour Agahi, Negar Akasheh, Afshin Ahmadvand, Hossein Akbari, Fatemeh Izadpanah
    Diabetes & Metabolic Syndrome: Clinical Research &.2018; 12(1): 9.     CrossRef
  • Dengue viruses and promising envelope protein domain III-based vaccines
    Hossein Fahimi, Mahshid Mohammadipour, Hamed Haddad Kashani, Farshid Parvini, Majid Sadeghizadeh
    Applied Microbiology and Biotechnology.2018; 102(7): 2977.     CrossRef
  • Recombinant Endolysins as Potential Therapeutics against Antibiotic-Resistant Staphylococcus aureus: Current Status of Research and Novel Delivery Strategies
    Hamed Haddad Kashani, Mathias Schmelcher, Hamed Sabzalipoor, Elahe Seyed Hosseini, Rezvan Moniri
    Clinical Microbiology Reviews.2018;[Epub]     CrossRef
  • A Novel Chimeric Endolysin with Antibacterial Activity against Methicillin-Resistant Staphylococcus aureus
    Hamed Haddad Kashani, Hossein Fahimi, Yasaman Dasteh Goli, Rezvan Moniri
    Frontiers in Cellular and Infection Microbiology.2017;[Epub]     CrossRef
  • The lowering of bilirubin levels in patients with neonatal jaundice using massage therapy: A randomized, double-blind clinical trial
    Fatemeh Eghbalian, Haneyeh Rafienezhad, Javad Farmal
    Infant Behavior and Development.2017; 49: 31.     CrossRef
  • The effect of chronic noise stress on serum levels of cortisol, gonadotropins, and sexual hormones at implantation time of mice
    Alireza Shafiei, Hassan Ehteram, Hossein Akbari, Masoud Motalebi Kashani, Mandana Beigi, Javad Amini Mahabadi, Tahere Mazoochi
    Comparative Clinical Pathology.2017; 26(4): 779.     CrossRef
  • The Role of Probiotics in the Treatment of Dysentery: a Randomized Double-Blind Clinical Trial
    Alireza Sharif, Hamed Haddad Kashani, Elahe Nasri, Zahra Soleimani, Mohammad Reza Sharif
    Probiotics and Antimicrobial Proteins.2017; 9(4): 380.     CrossRef
  • Assessment of 25-hydroxy vitamin D in stroke patients based on severity and type: a cross-sectional study
    Ebrahim Kouchaki, Mansour Sayyah, Maryam Omidvari
    Comparative Clinical Pathology.2017; 26(4): 811.     CrossRef
  • Purification of Antibacterial CHAPK Protein Using a Self-Cleaving Fusion Tag and Its Activity Against Methicillin-Resistant Staphylococcus aureus
    Elahe Seyed Hosseini, Rezvan Moniri, Yasaman Dasteh Goli, Hamed Haddad Kashani
    Probiotics and Antimicrobial Proteins.2016; 8(4): 202.     CrossRef

PHRP : Osong Public Health and Research Perspectives