Cloning, expression, purification and functional analysis of a specific multi-epitope protein from multi drug resistance Acinetobacter baumannii
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Abstract
Background and Objectives: Immunization is a promising strategy to combat against the life-threatening infections by Multi Drug Resistance Acinetobacter baumannii. In this study, we directed to design and evaluate the efficacy of a recombinant multi-epitope protein against this pathogen.
Materials and Methods: Epitopes prediction was performed for candidate proteins OmpA and BAM complex (BamA, BamB, BamC, BamD, BamE) from A. baumannii, using immune-informatics tools with high affinity for the human HLA alleles. After expression and purification of the recombinant protein, its functional activity was confirmed by interaction with positive sera.
Results: Cloning and expression of the desired multi-epitopes protein were verified. Circular Dichroism study showed the secondary structure and proper refolding of the recombinant protein was achieved and matched with computational prediction. There was a significant interaction between designed protein with antibodies presented in ICU patients' and staff's sera.
Conclusion: The interaction of the recombinant protein with patients' sera antibodies suggests that it may be a promising determinant protein for immunization against of MDR A. baumannii.
1. Prashanth K, Badrinath S. Nosocomial infections due to Acinetobacter species: clinical findings, risk and prognostic factors. Indian J Med Microbiol 2006; 24: 39-44.
2. Peleg AY, Seifert H, Paterson DL. Acinetobacter baumannii: emergence of a successful pathogen. Clin Microbiol Rev 2008; 21: 538-582.
3. Shields RK, Clancy CJ, Gillis LM, Kwak EJ, Silveira FP, Massih RC, et al. Epidemiology, clinical characteristics and outcomes of extensively drug-resistant Acinetobacter baumannii infections among solid organ transplant recipients. PLoS One 2012; 7(12): e52349.
4. Perez F, Hujer AM, Hujer KM, Decker BK, Rather PN, Bonomo RA. Global challenge of multidrug-resistant Acinetobacter baumannii. Antimicrob Agents Chemother 2007; 51: 3471-3484.
5. Mishra RP, Oviedo-Orta E, Prachi P, Rappuoli R, Bagnoli F. Vaccines and antibiotic resistance. Curr Opin Microbiol 2012; 15: 596-602.
6. Bentancor LV, Routray A, Bozkurt-Guzel C, Camacho-Peiro A, Pier GB, Maira-Litrán T. Evaluation of the trimeric autotransporter Ata as a vaccine candidate against Acinetobacter baumannii infections. Infect Immun 2012; 80: 3381-3388.
7. Golshani M, Rafati S, Jahanian-Najafabadi A, Nejati-Moheimani M, Siadat SD, Shahcheraghi F, et al. In silico design, cloning and high level expression of L7/L12-TOmp31 fusion protein of Brucella antigens. Res Pharm Sci 2015; 10: 436-445.
8. Singh R, Garg N, Shukla G, Capalash N, Sharma P. Immunoprotective efficacy of Acinetobacter baumannii outer membrane protein, FilF, predicted in silico as a potential vaccine candidate. Front Microbiol 2016; 7: 158.
9. Russo TA, Beanan JM, Olson R, MacDonald U, Cox AD, St Michael F, et al. The K1 capsular polysaccharide from Acinetobacter baumannii is a potential therapeutic target via passive immunization. Infect Immun 2013; 81: 915-922.
10. Noori E, Rasooli I, Owlia P, Mousavi Gargari SL, Ebrahimizadeh W. A conserved region from biofilm associated protein as a biomarker for detection of Acinetobacter baumannii. Microb Pathog 2014; 77: 84-88.
11. McConnell MJ, Rumbo C, Bou G, Pachón J. Outer membrane vesicles as an acellular vaccine against Acinetobacter baumannii. Vaccine 2011; 29: 5705-5710.
12. Nie D, Hu Y, Chen Z, Li M, Hou Z, Luo X, et al. Outer membrane protein A (OmpA) as a potential therapeutic target for Acinetobacter baumannii infection. J Biomed Sci 2020; 27: 26.|
13. Rasooli I, Abdolhamidi R, Jahangiri A, Darvish Alipour Astaneh S. Outer membrane protein, Oma87 prevents Acinetobacter baumannii infection. Int J Pept Res Ther 2020; 1-8.
14. Huang W, Yao Y, Wang S, Xia Y, Yang X, Long Q, et al. Immunization with a 22-kDa outer membrane protein elicits protective immunity to multidrug-resistant Acinetobacter baumannii. Sci Rep 2016; 6: 20724.
15. Gaddy JA, Tomaras AP, Actis LA. The Acinetobacter baumannii 19606 OmpA protein plays a role in biofilm formation on abiotic surfaces and in the interaction of this pathogen with eukaryotic cells. Infect Immun 2009; 77: 3150-3160.
16. Moon DC, Choi CH, Lee JH, Choi CW, Kim HY, Park JS, et al. Acinetobacter baumannii outer membrane protein A modulates the biogenesis of outer membrane vesicles. J Microbiol 2012; 50: 155-160.
17. Choi CH, Lee EY, Lee YC, Park TI, Kim HJ, Hyun SH, et al. Outer membrane protein 38 of Acinetobacter baumannii localizes to the mitochondria and induces apoptosis of epithelial cells. Cell Microbiol 2005; 7: 1127-1138.
18. Hagan CL, Wzorek JS, Kahne D. Inhibition of the β-barrel assembly machine by a peptide that binds BamD. Proc Natl Acad Sci U S A 2015; 112: 2011-2016.
19. Hagan CL, Silhavy TJ, Kahne D. β-Barrel membrane protein assembly by the Bam complex. Annu Rev Biochem 2011; 80: 189-210.
20. Knowles TJ, Scott-Tucker A, Overduin M, Henderson IR. Membrane protein architects: the role of the BAM complex in outer membrane protein assembly. Nat Rev Microbiol 2009; 7: 206-214.
21. Guo SJ, Ren S, Xie YE. Evaluation of the protective efficacy of a fused OmpK/Omp22 protein vaccine candidate against Acinetobacter baumannii infection in mice. Biomed Environ Sci 2018; 31: 155-158.
22. Ren S, Guan L, Dong Y, Wang C, Feng L, Xie Y. Design and evaluation of a multi-epitope assembly peptide vaccine against Acinetobacter baumannii infection in mice. Swiss Med Wkly 2019; 149: w20052.
23. Badmasti F, Ajdary S, Bouzari S, Fooladi AA, Shahcheraghi F, Siadat SD. Immunological evaluation of OMV (PagL)+ Bap (1-487aa) and AbOmpA (8-346aa)+ Bap (1-487aa) as vaccine candidates against Acinetobacter baumannii sepsis infection. Mol Immunol 2015; 67(2 Pt B): 552-558.
24. Bambini S, Rappuoli R. The use of genomics in microbial vaccine development. Drug Discov Today 2009; 14: 252-260.
25. Soria-Guerra RE, Nieto-Gomez R, Govea-Alonso DO, Rosales-Mendoza S. An overview of bioinformatics tools for epitope prediction: implications on vaccine development. J Biomed Inform 2015; 53: 405-414.
26. Mohammadi F, Goudarzi H, Hashemi A, Yousefi Nojookambari N, Khoshnood S, Sabzehali F. Detection of ISAba1 in Acinetobacter baumannii strains carrying OXA genes isolated from Iranian burns patients. Arch Pediatr Infect Dis 2016; 5(2); e39307.
27. Nafarieh T, Bandehpour M, Hashemi A, Taheri S, Yardel V, Jamaati H, et al. Identification of antigens from nosocomial Acinetobacter baumannii clinical isolates in sera from ICU staff and infected patients using the antigenome technique. World J Microbiol Biotechnol 2017; 33: 189.
28. Jositsch G, Papadakis T, Haberberger RV, Wolff M, Wess J, Kummer W. Suitability of muscarinic acetylcholine receptor antibodies for immunohistochemistry evaluated on tissue sections of receptor gene-deficient mice. Naunyn Schmiedebergs Arch Pharmacol 2009; 379: 389-395.
29 Ho BK, Brasseur R. The Ramachandran plots of glycine and pre-proline. BMC Struct Biol 2005; 5: 14.
30. Xie R, Zhang XD, Zhao Q, Peng B, Zheng J. Analysis of global prevalence of antibiotic resistance in Acinetobacter baumannii infections disclosed a faster increase in OECD countries. Emerg Microbes Infect 2018; 7: 31.
31. Chiang MH, Sung WC, Lien SP, Chen YZ, Lo AF, Huang JH, et al. Identification of novel vaccine candidates against Acinetobacter baumannii using reverse vaccinology. Hum vaccin Immunother 2015; 11: 1065-1073.
32. Gomez G, Pei J, Mwangi W, Adams LG, Rice-Ficht A, Ficht TA. Immunogenic and invasive properties of Brucella melitensis 16M outer membrane protein vaccine candidates identified via a reverse vaccinology approach. PLoS One 2013; 8(3): e59751.
33. Ahmadi K, Pouladfar G, Kalani M, Faezi S, Pourmand MR, Hasanzadeh S, et al. Epitope-based immunoinformatics study of a novel Hla-MntC-SACOL0723 fusion protein from Staphylococcus aureus: induction of multi-pattern immune responses. Mol Immunol 2019; 114: 88-99.
34. Esmailnia E, Amani J, Gargari SLM. Identification of novel vaccine candidate against Salmonella enterica serovar Typhi by reverse vaccinology method and evaluation of its immunization. Genomics 2020; 112: 3374-3381.
35. Sadeghi Z, Fasihi-Ramandi M, Bouzari S. Evaluation of immunogenicity of novel multi-epitope subunit vaccines in combination with poly I:C against Brucella melitensis and Brucella abortus infection. Int Immunopharmacol 2019; 75: 105829.
36. Chopra S, Ramkissoon K, Anderson DC. A systematic quantitative proteomic examination of multidrug resistance in Acinetobacter baumannii. J Proteomics 2013; 84: 17-39.
37. Lin L, Tan B, Pantapalangkoor P, Ho T, Hujer AM, Taracila MA, et al. Acinetobacter baumannii rOmpA vaccine dose alters immune polarization and immunodominant epitopes. Vaccine 2013; 31: 313-318.
38. Fajardo Bonin R, Chapeaurouge A, Perales J, da Silva JG Jr, do Nascimento HJ, D'Alincourt Carvalho, et al. Identification of immunogenic proteins of the bacterium Acinetobacter baumannii using a proteomic approach. Proteomics Clin Appl 2014; 8: 916-923.
39. Ansari H, Doosti A, Kargar M, Bijanzadeh M, Jaafarinia M. Cloning of ompA gene from Acinetobacter baumannii into the eukaryotic expression vector pBudCE4.1 as DNA vaccine. Indian J Microbiol 2018; 58: 174-181.
40. Bolourchi N, Shahcheraghi F, Shirazi AS, Janani A, Bahrami F, Badmasti F. Immunogenic reactivity of recombinant PKF and AbOmpA proteins as serum resistance factors against sepsis of Acinetobacter baumannii. Microb Pathog 2019; 131: 9-14.
41. Singh R, Capalash N, Sharma P. Immunoprotective potential of BamA, the outer membrane protein assembly factor, against MDR Acinetobacter baumannii. Sci Rep 2017; 7: 12411.
42. Yari F, Sobhani M, Sabaghi F, Zaman-Vaziri M, Bagheri N, Talebian A. Frequencies of HLA-DRB1 in Iranian normal population and in patients with acute lymphoblastic leukemia. Arch Med Res 2008; 39: 205-208.
43. Chen X, Zaro JL, Shen WC. Fusion protein linkers: property, design and functionality. Adv Drug Deliv Rev 2013; 65: 1357-1369.
44. Mehdinejadiani K, Bandehpour M, Hashemi A, Ranjbar MM, Taheri S, Jalali SA, et al. In silico design and evaluation of Acinetobacter baumannii outer membrane protein A (OmpA) antigenic peptides as vaccine candidate in immunized mice. Iran J Allergy Asthma Immunol 2019; 18: 655-663.
45. Ghazi FMP, Gargari SLM. Synthetic peptides mimicking lipopolysaccharide as a potential vaccine candidate against Vibrio cholerae serogroup O1. Iran J Microbiol 2017; 9: 244-250.
2. Peleg AY, Seifert H, Paterson DL. Acinetobacter baumannii: emergence of a successful pathogen. Clin Microbiol Rev 2008; 21: 538-582.
3. Shields RK, Clancy CJ, Gillis LM, Kwak EJ, Silveira FP, Massih RC, et al. Epidemiology, clinical characteristics and outcomes of extensively drug-resistant Acinetobacter baumannii infections among solid organ transplant recipients. PLoS One 2012; 7(12): e52349.
4. Perez F, Hujer AM, Hujer KM, Decker BK, Rather PN, Bonomo RA. Global challenge of multidrug-resistant Acinetobacter baumannii. Antimicrob Agents Chemother 2007; 51: 3471-3484.
5. Mishra RP, Oviedo-Orta E, Prachi P, Rappuoli R, Bagnoli F. Vaccines and antibiotic resistance. Curr Opin Microbiol 2012; 15: 596-602.
6. Bentancor LV, Routray A, Bozkurt-Guzel C, Camacho-Peiro A, Pier GB, Maira-Litrán T. Evaluation of the trimeric autotransporter Ata as a vaccine candidate against Acinetobacter baumannii infections. Infect Immun 2012; 80: 3381-3388.
7. Golshani M, Rafati S, Jahanian-Najafabadi A, Nejati-Moheimani M, Siadat SD, Shahcheraghi F, et al. In silico design, cloning and high level expression of L7/L12-TOmp31 fusion protein of Brucella antigens. Res Pharm Sci 2015; 10: 436-445.
8. Singh R, Garg N, Shukla G, Capalash N, Sharma P. Immunoprotective efficacy of Acinetobacter baumannii outer membrane protein, FilF, predicted in silico as a potential vaccine candidate. Front Microbiol 2016; 7: 158.
9. Russo TA, Beanan JM, Olson R, MacDonald U, Cox AD, St Michael F, et al. The K1 capsular polysaccharide from Acinetobacter baumannii is a potential therapeutic target via passive immunization. Infect Immun 2013; 81: 915-922.
10. Noori E, Rasooli I, Owlia P, Mousavi Gargari SL, Ebrahimizadeh W. A conserved region from biofilm associated protein as a biomarker for detection of Acinetobacter baumannii. Microb Pathog 2014; 77: 84-88.
11. McConnell MJ, Rumbo C, Bou G, Pachón J. Outer membrane vesicles as an acellular vaccine against Acinetobacter baumannii. Vaccine 2011; 29: 5705-5710.
12. Nie D, Hu Y, Chen Z, Li M, Hou Z, Luo X, et al. Outer membrane protein A (OmpA) as a potential therapeutic target for Acinetobacter baumannii infection. J Biomed Sci 2020; 27: 26.|
13. Rasooli I, Abdolhamidi R, Jahangiri A, Darvish Alipour Astaneh S. Outer membrane protein, Oma87 prevents Acinetobacter baumannii infection. Int J Pept Res Ther 2020; 1-8.
14. Huang W, Yao Y, Wang S, Xia Y, Yang X, Long Q, et al. Immunization with a 22-kDa outer membrane protein elicits protective immunity to multidrug-resistant Acinetobacter baumannii. Sci Rep 2016; 6: 20724.
15. Gaddy JA, Tomaras AP, Actis LA. The Acinetobacter baumannii 19606 OmpA protein plays a role in biofilm formation on abiotic surfaces and in the interaction of this pathogen with eukaryotic cells. Infect Immun 2009; 77: 3150-3160.
16. Moon DC, Choi CH, Lee JH, Choi CW, Kim HY, Park JS, et al. Acinetobacter baumannii outer membrane protein A modulates the biogenesis of outer membrane vesicles. J Microbiol 2012; 50: 155-160.
17. Choi CH, Lee EY, Lee YC, Park TI, Kim HJ, Hyun SH, et al. Outer membrane protein 38 of Acinetobacter baumannii localizes to the mitochondria and induces apoptosis of epithelial cells. Cell Microbiol 2005; 7: 1127-1138.
18. Hagan CL, Wzorek JS, Kahne D. Inhibition of the β-barrel assembly machine by a peptide that binds BamD. Proc Natl Acad Sci U S A 2015; 112: 2011-2016.
19. Hagan CL, Silhavy TJ, Kahne D. β-Barrel membrane protein assembly by the Bam complex. Annu Rev Biochem 2011; 80: 189-210.
20. Knowles TJ, Scott-Tucker A, Overduin M, Henderson IR. Membrane protein architects: the role of the BAM complex in outer membrane protein assembly. Nat Rev Microbiol 2009; 7: 206-214.
21. Guo SJ, Ren S, Xie YE. Evaluation of the protective efficacy of a fused OmpK/Omp22 protein vaccine candidate against Acinetobacter baumannii infection in mice. Biomed Environ Sci 2018; 31: 155-158.
22. Ren S, Guan L, Dong Y, Wang C, Feng L, Xie Y. Design and evaluation of a multi-epitope assembly peptide vaccine against Acinetobacter baumannii infection in mice. Swiss Med Wkly 2019; 149: w20052.
23. Badmasti F, Ajdary S, Bouzari S, Fooladi AA, Shahcheraghi F, Siadat SD. Immunological evaluation of OMV (PagL)+ Bap (1-487aa) and AbOmpA (8-346aa)+ Bap (1-487aa) as vaccine candidates against Acinetobacter baumannii sepsis infection. Mol Immunol 2015; 67(2 Pt B): 552-558.
24. Bambini S, Rappuoli R. The use of genomics in microbial vaccine development. Drug Discov Today 2009; 14: 252-260.
25. Soria-Guerra RE, Nieto-Gomez R, Govea-Alonso DO, Rosales-Mendoza S. An overview of bioinformatics tools for epitope prediction: implications on vaccine development. J Biomed Inform 2015; 53: 405-414.
26. Mohammadi F, Goudarzi H, Hashemi A, Yousefi Nojookambari N, Khoshnood S, Sabzehali F. Detection of ISAba1 in Acinetobacter baumannii strains carrying OXA genes isolated from Iranian burns patients. Arch Pediatr Infect Dis 2016; 5(2); e39307.
27. Nafarieh T, Bandehpour M, Hashemi A, Taheri S, Yardel V, Jamaati H, et al. Identification of antigens from nosocomial Acinetobacter baumannii clinical isolates in sera from ICU staff and infected patients using the antigenome technique. World J Microbiol Biotechnol 2017; 33: 189.
28. Jositsch G, Papadakis T, Haberberger RV, Wolff M, Wess J, Kummer W. Suitability of muscarinic acetylcholine receptor antibodies for immunohistochemistry evaluated on tissue sections of receptor gene-deficient mice. Naunyn Schmiedebergs Arch Pharmacol 2009; 379: 389-395.
29 Ho BK, Brasseur R. The Ramachandran plots of glycine and pre-proline. BMC Struct Biol 2005; 5: 14.
30. Xie R, Zhang XD, Zhao Q, Peng B, Zheng J. Analysis of global prevalence of antibiotic resistance in Acinetobacter baumannii infections disclosed a faster increase in OECD countries. Emerg Microbes Infect 2018; 7: 31.
31. Chiang MH, Sung WC, Lien SP, Chen YZ, Lo AF, Huang JH, et al. Identification of novel vaccine candidates against Acinetobacter baumannii using reverse vaccinology. Hum vaccin Immunother 2015; 11: 1065-1073.
32. Gomez G, Pei J, Mwangi W, Adams LG, Rice-Ficht A, Ficht TA. Immunogenic and invasive properties of Brucella melitensis 16M outer membrane protein vaccine candidates identified via a reverse vaccinology approach. PLoS One 2013; 8(3): e59751.
33. Ahmadi K, Pouladfar G, Kalani M, Faezi S, Pourmand MR, Hasanzadeh S, et al. Epitope-based immunoinformatics study of a novel Hla-MntC-SACOL0723 fusion protein from Staphylococcus aureus: induction of multi-pattern immune responses. Mol Immunol 2019; 114: 88-99.
34. Esmailnia E, Amani J, Gargari SLM. Identification of novel vaccine candidate against Salmonella enterica serovar Typhi by reverse vaccinology method and evaluation of its immunization. Genomics 2020; 112: 3374-3381.
35. Sadeghi Z, Fasihi-Ramandi M, Bouzari S. Evaluation of immunogenicity of novel multi-epitope subunit vaccines in combination with poly I:C against Brucella melitensis and Brucella abortus infection. Int Immunopharmacol 2019; 75: 105829.
36. Chopra S, Ramkissoon K, Anderson DC. A systematic quantitative proteomic examination of multidrug resistance in Acinetobacter baumannii. J Proteomics 2013; 84: 17-39.
37. Lin L, Tan B, Pantapalangkoor P, Ho T, Hujer AM, Taracila MA, et al. Acinetobacter baumannii rOmpA vaccine dose alters immune polarization and immunodominant epitopes. Vaccine 2013; 31: 313-318.
38. Fajardo Bonin R, Chapeaurouge A, Perales J, da Silva JG Jr, do Nascimento HJ, D'Alincourt Carvalho, et al. Identification of immunogenic proteins of the bacterium Acinetobacter baumannii using a proteomic approach. Proteomics Clin Appl 2014; 8: 916-923.
39. Ansari H, Doosti A, Kargar M, Bijanzadeh M, Jaafarinia M. Cloning of ompA gene from Acinetobacter baumannii into the eukaryotic expression vector pBudCE4.1 as DNA vaccine. Indian J Microbiol 2018; 58: 174-181.
40. Bolourchi N, Shahcheraghi F, Shirazi AS, Janani A, Bahrami F, Badmasti F. Immunogenic reactivity of recombinant PKF and AbOmpA proteins as serum resistance factors against sepsis of Acinetobacter baumannii. Microb Pathog 2019; 131: 9-14.
41. Singh R, Capalash N, Sharma P. Immunoprotective potential of BamA, the outer membrane protein assembly factor, against MDR Acinetobacter baumannii. Sci Rep 2017; 7: 12411.
42. Yari F, Sobhani M, Sabaghi F, Zaman-Vaziri M, Bagheri N, Talebian A. Frequencies of HLA-DRB1 in Iranian normal population and in patients with acute lymphoblastic leukemia. Arch Med Res 2008; 39: 205-208.
43. Chen X, Zaro JL, Shen WC. Fusion protein linkers: property, design and functionality. Adv Drug Deliv Rev 2013; 65: 1357-1369.
44. Mehdinejadiani K, Bandehpour M, Hashemi A, Ranjbar MM, Taheri S, Jalali SA, et al. In silico design and evaluation of Acinetobacter baumannii outer membrane protein A (OmpA) antigenic peptides as vaccine candidate in immunized mice. Iran J Allergy Asthma Immunol 2019; 18: 655-663.
45. Ghazi FMP, Gargari SLM. Synthetic peptides mimicking lipopolysaccharide as a potential vaccine candidate against Vibrio cholerae serogroup O1. Iran J Microbiol 2017; 9: 244-250.
| Files | ||
| Issue | Vol 13 No 5 (2021) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v13i5.7429 | |
| Keywords | ||
| Multi drug resistant; Acinetobacter baumannii; Recombinant multi-epitope protein (rMEP); OmpA; BAM complex | ||
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This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
How to Cite
1.
Davoudi Z, Taromchi A, Kazemi B, Bandehpour M, Mosaffa N. Cloning, expression, purification and functional analysis of a specific multi-epitope protein from multi drug resistance Acinetobacter baumannii. Iran J Microbiol. 2021;13(5):642-652.
