KaydedilenlerDerginizi ekleyin

Dergiler / Osmaniye Korkut Ata Üniversitesi Fen Bilimleri Enstitüsü Dergisi / 2026 / Cilt: 6 Sayı: 2

Escherichia coli'de Acinetobacter baumannii OccAB1 Porin Proteininin Klonlanması, Ekspresyonu ve Saflaştırılması

Cloning, Expression, and Purification of Acinetobacter baumannii OccAB1 Porin Protein in Escherichia coli

Özge Kaygusuz İzgördü(BİLECİK ŞEYH EDEBALİ ÜNİVERSİTESİ)
Cihan Darcan(BILECIK SEYH EDEBALI UNIVERSITY)0000-0003-0205-3774
Fatma Doğan Guzel(YILDIRIM BEYAZIT UNIVERSITY)
Sayfa
1529–1542
Yayın Tarihi
2023-07-05
DOI
—
🏛️Bu makalenin bibliyografik verileri ve tam metin bağlantısı DergiPark üzerinden sağlanmıştır.
DergiPark Kaydı ↗
✨

Yapay Zeka Hızlı Bakış & Çıkarımlar

AI Insight
⚡ TL;DR(1 Cümlede Hızlı Çıkarım)

Porinler ve dışa akış pompalarının aşırı ekspresyonu, ilaç direncini artırmada ve tedavinin etkisiz hale getirilmesinde katkıda bulunan faktörler olarak tespit edilmiştir.

📌 3 Maddede Bu Makale

🎯
Amaç & Hipotez:Acinetobacter baumannii, toprakta, suda, insan ve hayvan vücutlarının yüzeyleri gibi farklı habitatlarda serbest yaşayan gram negatif bir organizmalardır.
📊
Temel Bulgular & Veri:Porinler ve dışa akış pompalarının aşırı ekspresyonu, ilaç direncini artırmada ve tedavinin etkisiz hale getirilmesinde katkıda bulunan faktörler olarak tespit edilmiştir.
💡
Sonuç & Katkı:Sonuç olarak, ekspresyon vektörüne klonlanan A. baumannii OccAB1 porini, E. coli'de başarıyla eksprese edildi.
🔬Tür:Araştırma Makalesi
📍Kapsam:Genişletilmiş Akademik Örneklem
⚙️Değişkenler:E. coli · A. baumannii · Ni-NTA

Özet

Acinetobacter baumannii, toprakta, suda, insan ve hayvan vücutlarının yüzeyleri gibi farklı habitatlarda serbest yaşayan gram negatif bir organizmalardır. Ancak zamanla patojenik bir yaşam tarzına adapte olarak tüm dünyada yoğun bakım ünitelerinde enfeksiyonlara ve ölümlere neden olmaktadır. Antibiyotik direncinde porin proteinleri bu bakteride temel rol oynamaktadır. Porinler ve dışa akış pompalarının aşırı ekspresyonu, ilaç direncini artırmada ve tedavinin etkisiz hale getirilmesinde katkıda bulunan faktörler olarak tespit edilmiştir. Bu sebeple antibiyotik geçişlerinin in vitro olarak çalışılabilmesi için öncelikli olarak bu proteinlerin saflaştırılması gerekmektedir. Saflaştırma işleminin ardından elde edilen bu porin proteini ile çift katmanlı lipit tabakalarda antibiyotiklerin geçişleri araştırılacaktır. Bu çalışmada, A. baumannii OccAB1 porin proteini klonlandı ve E. coli'de eksprese edildi. Sonuç olarak, ekspresyon vektörüne klonlanan A. baumannii OccAB1 porini, E. coli'de başarıyla eksprese edildi.

Abstract

Acinetobacter baumannii is a free-living gram-negative organism in different habitats such as soil, water, surfaces of human and animal bodies. However, adapting to a pathogenic lifestyle over time, it causes infections and deaths in intensive care units all over the world. Porin proteins play a fundamental role in antibiotic resistance in this bacterium. Overexpression of porins and efflux pumps have been identified as contributing factors in increasing drug resistance and inactivation of therapy. For this reason, to study antibiotic transitions in vitro, these proteins must be purified first. With this porin protein obtained after purification, the passage of antibiotics in bilayer lipid layers will be investigated. In this study, A. baumannii OccAB1 porin protein was cloned and expressed in E. coli. As a result, A. baumannii OccAB1 porin cloned into the expression vector was successfully expressed in E. coli.

🔖

Bu Makaleye Atıf Yap

İzgördü, Ö. K., Darcan, C., Guzel, F. D. (2023). Escherichia coli'de Acinetobacter baumannii OccAB1 Porin Proteininin Klonlanması, Ekspresyonu ve Saflaştırılması. Osmaniye Korkut Ata Üniversitesi Fen Bilimleri Enstitüsü Dergisi, Cilt: 6 Sayı: 2, 1529–1542. https://ajindex.com/articles/1534360

📄 Tam Metin PDF Okuyucu

📄

PDF görüntüleyici hazırlanıyor...

Kaynakça (45)

  1. Abbott, I., Cerqueira, G. M., Bhuiyan, S., & Peleg, A. Y. (2013). Carbapenem resistance in Acinetobacter baumannii: laboratory challenges, mechanistic insights, and therapeutic strategies. Expert Review of Anti-Infective Therapy, 11(4), 395–409. https://doi.org/10.1586/ERI.13.21
  2. Assenberg, R., Wan, P. T., Geisse, S., & Mayr, L. M. (2013). Advances in recombinant protein expression for use in pharmaceutical research. Current Opinion in Structural Biology, 23(3), 393–402. https://doi.org/10.1016/J.SBI.2013.03.008
  3. Bhamidimarri, S. P., Zahn, M., Prajapati, J. D., Schleberger, C., Söderholm, S., Hoover, J., West, J., Kleinekathöfer, U., Bumann, D., Winterhalter, M., & van den Berg, B. (2019). A Multidisciplinary Approach toward Identification of Antibiotic Scaffolds for Acinetobacter baumannii. Structure (London, England: 1993), 27(2), 268-280.e6. https://doi.org/10.1016/J.STR.2018.10.021
  4. Blattner, F. R., Plunkett, G., Bloch, C. A., Perna, N. T., Burland, V., Riley, M., Collado-Vides, J., Glasner, J. D., Rode, C. K., Mayhew, G. F., Gregor, J., Davis, N. W., Kirkpatrick, H. A., Goeden, M. A., Rose, D. J., Mau, B., & Shao, Y. (1997). The complete genome sequence of Escherichia coli K-12. Science (New York, N.Y.), 277(5331), 1453–1462. https://doi.org/10.1126/SCIENCE.277.5331.1453
  5. Borneleit, P., & Kleber, H.-P. (1991). The Outer Membrane of: Structure-Function Relationships. The Biology of Acinetobacter, 259–271. https://doi.org/10.1007/978-1-4899-3553-3_18
  6. Choi, U., & Lee, C. R. (2019). Distinct Roles of Outer Membrane Porins in Antibiotic Resistance and Membrane Integrity in Escherichia coli. Frontiers in Microbiology, 10(APR), 1–9. https://doi.org/10.3389/fmicb.2019.00953
  7. Clark, T. J., Momany, C., & Neidle, E. L. (2002). The benPK operon, proposed to play a role in transport, is part of a regulon for benzoate catabolism in Acinetobacter sp. strain ADP1. Microbiology (Reading, England), 148(Pt 4), 1213–1223. https://doi.org/10.1099/00221287-148-4-1213 Darcan, C., Özkanca, R., İdil. Ö. (2009). The role of RpoS, H-NS and AcP on the pH-dependent OmpC and OmpF porin expressions of Escherichia coli at different pH, Afrıcan Journal of Bıotechnology 8(9):1845-1854.
  8. Darcan, C. (2012). Expression of OmpC and OmpF porin proteins and survival of escherichia coli under photooxidative stress in black seawater. Aquatic Biology, 17(2), 97–105. https://doi.org/10.3354/AB00458
  9. Darcan, C., & Aydin, E. (2012). Fur- mutation increases the survival time of Escherichia coli under photooxidative stress in aquatic environments. Acta Biologica Hungarica, 63(3), 399–409. https://doi.org/10.1556/ABIOL.63.2012.3.10
  10. Darcan, C., Özkanca, R., & Flint, K. P. (2003). Survival of nonspecific porin-deficient mutants of Escherichia coli in black seawater. Letters in Applied Microbiology, 37(5), 380–385. https://doi.org/10.1046/j.1472-765X.2003.01418.x
  11. Demain, A. L., & Vaishnav, P. (2009). Production of recombinant proteins by microbes and higher organisms. Biotechnology Advances, 27(3), 297–306. https://doi.org/10.1016/J.BIOTECHADV.2009.01.008
  12. Dupont, M., Pagès, J. M., Lafitte, D., Siroy, A., & Bollet, C. (2005). Identification of an OprD homologue in Acinetobacter baumannii. Journal of Proteome Research, 4(6), 2386–2390. https://doi.org/10.1021/PR050143Q/ASSET/IMAGES/PR050143Q.SOCIAL.JPEG_V03
  13. Esterly, J. S., Griffith, M., Qi, C., Malczynski, M., Postelnick, M. J., & Scheetz, M. H. (2011). Impact of carbapenem resistance and receipt of active antimicrobial therapy on clinical outcomes of Acinetobacter baumannii bloodstream infections. Antimicrobial Agents and Chemotherapy, 55(10), 4844–4849. https://doi.org/10.1128/AAC.01728-10
  14. He, S., Wu, X., Ma, B., & Xu, Y. (2021). High specific immobilization of His-tagged recombinant Microbacterium esterase by Ni-NTA magnetic chitosan microspheres for efficient synthesis of key chiral intermediate of d-biotin. Bioprocess and Biosystems Engineering, 44(10), 2193–2204. https://doi.org/10.1007/S00449-021-02595-7
  15. Huang, C. J., Lin, H., & Yang, X. (2012). Industrial production of recombinant therapeutics in Escherichia coli and its recent advancements. Journal of Industrial Microbiology and Biotechnology, 39(3), 383–399. https://doi.org/10.1007/S10295-011-1082-9
  16. Joshi, H., & Jain, V. (2017). Novel method to rapidly and efficiently lyse Escherichia coli for the isolation of recombinant protein. Analytical Biochemistry, 528, 1–6. https://doi.org/10.1016/J.AB.2017.04.009
  17. Lee, C. R., Cho, I. H., Jeong, B. C., & Lee, S. H. (2013). Strategies to Minimize Antibiotic Resistance. International Journal of Environmental Research and Public Health 2013, Vol. 10, Pages 4274-4305, 10(9), 4274–4305. https://doi.org/10.3390/IJERPH10094274
  18. Lee, C. R., Lee, J. H., Park, M., Park, K. S., Bae, I. K., Kim, Y. B., Cha, C. J., Jeong, B. C., & Lee, S. H. (2017). Biology of Acinetobacter baumannii: Pathogenesis, Antibiotic Resistance Mechanisms, and Prospective Treatment Options. Frontiers in Cellular and Infection Microbiology, 7(MAR). https://doi.org/10.3389/FCIMB.2017.00055
  19. Mostachio, A. K., Levin, A. S., Rizek, C., Rossi, F., Zerbini, J., & Costa, S. F. (2012). High prevalence of OXA-143 and alteration of outer membrane proteins in carbapenem-resistant Acinetobacter spp. isolates in Brazil. International Journal of Antimicrobial Agents, 39(5), 396–401. https://doi.org/10.1016/J.IJANTIMICAG.2012.01.021
  20. Nikaido, H. (2003). Molecular Basis of Bacterial Outer Membrane Permeability Revisited. Microbiology and Molecular Biology Reviews, 67(4), 593–656. https://doi.org/10.1128/MMBR.67.4.593-656.2003/FORMAT/EPUB
  21. Nikaido, H., & Vaara, M. (1985). Molecular basis of bacterial outer membrane permeability. Microbiological Reviews, 49(1), 1–32. https://doi.org/10.1128/mmbr.49.1.1-32.1985
  22. Nurjayadi, M., Setiyoto, T., Jinan, S. F., Hardianto, D., Sulfianti, A., Agustini, K., & El-Enshasy, H. A. (2021). Purification of Fim-C-Salmonella typhi recombinant protein with Ni-NTA resin as raw material for typhoid disease detection kit. Journal of Physics: Conference Series, 1869(1). https://doi.org/10.1088/1742-6596/1869/1/012033
  23. Obara, M., & Nakae, T. (1991). Mechanisms of resistance to β-lactam antibiotics in Acinetobacter calcoaceticus. Journal of Antimicrobial Chemotherapy, 28(6), 791–800. https://doi.org/10.1093/JAC/28.6.791
  24. Overton, T. W. (2014). Recombinant protein production in bacterial hosts. Drug Discovery Today, 19(5), 590–601. https://doi.org/10.1016/j.drudis.2013.11.008
  25. Pagès, J. M., James, C. E., & Winterhalter, M. (2008). The porin and the permeating antibiotic: a selective diffusion barrier in Gram-negative bacteria. Nature Reviews Microbiology 2008 6:12, 6(12), 893–903. https://doi.org/10.1038/nrmicro1994
  26. Park, Y. K., Jung, S. I., Park, K. H., Kim, S. H., & Ko, K. S. (2012). Characteristics of carbapenem-resistant Acinetobacter spp. other than Acinetobacter baumannii in South Korea. International Journal of Antimicrobial Agents, 39(1), 81–85. https://doi.org/10.1016/J.IJANTIMICAG.2011.08.006
  27. Peränen, J., Rikkonen, M., Hyvönen, M., & Kääriäinen, L. (1996). T7 vectors with modified T7lac promoter for expression of proteins in Escherichia coli. Analytical Biochemistry, 236(2), 371–373. https://doi.org/10.1006/ABIO.1996.0187
  28. Rasooli, I., Abdolhamidi, R., Jahangiri, · Abolfazl, Darvish, S., & Astaneh, A. (2020). Outer Membrane Protein, Oma87 Prevents Acinetobacter baumannii Infection. International Journal of Peptide Research and Therapeutics, 26(3), 2653–2660. https://doi.org/10.1007/s10989-020-10056-0
  29. Rice, L. B. (2008). Federal Funding for the Study of Antimicrobial Resistance in Nosocomial Pathogens: No ESKAPE. The Journal of Infectious Diseases, 197(8), 1079–1081. https://doi.org/10.1086/533452
  30. Rice, L. B. (2010). Progress and challenges in implementing the research on ESKAPE pathogens. Infection Control and Hospital Epidemiology, 31 Suppl 1(S1), S7–S10. https://doi.org/10.1086/655995
  31. Santajit, S., & Indrawattana, N. (2016). Mechanisms of Antimicrobial Resistance in ESKAPE Pathogens. BioMed Research International, 2016. https://doi.org/10.1155/2016/2475067
  32. Segura, A., Bünz, P. v., D’Argenio, D. A., & Ornston, L. N. (1999). Genetic analysis of a chromosomal region containing vanA and vanB, genes required for conversion of either ferulate or vanillate to protocatechuate in Acinetobacter. Journal of Bacteriology, 181(11), 3494–3504. https://doi.org/10.1128/jb.181.11.3494-3504.1999
  33. Smith, M. A., Weaver, V. B., Young, D. M., & Ornston, L. N. (2003). Genes for chlorogenate and hydroxycinnamate catabolism (hca) are linked to functionally related genes in the dca-pca-qui-pob-hca chromosomal cluster of Acinetobacter sp. strain ADP1. Applied and Environmental Microbiology, 69(1), 524–532. https://doi.org/10.1128/AEM.69.1.524-532.2003
  34. Srinivasan, V. B., Vaidyanathan, V., & Rajamohan, G. (2015). AbuO, a tolc-like outer membrane protein of Acinetobacter baumannii, is involved in antimicrobial and oxidative stress resistance. Antimicrobial Agents and Chemotherapy, 59(2), 1236–1245. https://doi.org/10.1128/AAC.03626-14
  35. Studier, F. W. (1991). Use of bacteriophage T7 lysozyme to improve an inducible T7 expression system. Journal of Molecular Biology, 219(1), 37–44. https://doi.org/10.1016/0022-2836(91)90855-Z
  36. Sugawara, E., & Nikaido, H. (2012). OmpA is the principal nonspecific slow porin of Acinetobacter baumannii. Journal of Bacteriology, 194(15), 4089–4096. https://doi.org/10.1128/JB.00435-12
  37. Tokuda, H. (2014). Biogenesis of Outer Membranes in Gram-Negative Bacteria. OUP, 73(3), 465–473. https://doi.org/10.1271/BBB.80778 Uppalapati, S. R., Sett, A., & Pathania, R. (2020). The Outer Membrane Proteins OmpA, CarO, and OprD of Acinetobacter baumannii Confer a Two-Pronged Defense in Facilitating Its Success as a Potent Human Pathogen. Frontiers in Microbiology, 11, 2441. https://doi.org/10.3389/FMICB.2020.589234/BIBTEX
  38. Vashist, J., Tiwari, V., Kapil, A., & Rajeswari, M. R. (2010). Quantitative profiling and identification of outer membrane proteins of beta-lactam resistant strain of Acinetobacter baumannii. Journal of Proteome Research, 9(2), 1121–1128. https://doi.org/10.1021/PR9011188
  39. Vashistt, J., Tiwari, V., Kapil, A., & Moganty, R. R. (2011). Differential expression of outer membrane proteins in early stages of meropenem-resistance in Acinetobacter baumannii. Journal of Integrated OMICS, 1(2), 280. https://doi.org/10.5584/JIOMICS.V1I2.67
  40. Verma, S. K., Gautam, V., Balakrishna, K., & Kumar, S. (2009). Overexpression, purification, and immunogenicity of recombinant porin proteins of Salmonella enterica serovar Typhi (S. Typhi). Journal of Microbiology and Biotechnology, 19(9), 1034–1040. https://doi.org/10.4014/JMB.0812.675
  41. Vila, J., Martí, S., & Sánchez-Céspedes, J. (2007). Porins, efflux pumps and multidrug resistance in Acinetobacter baumannii. The Journal of Antimicrobial Chemotherapy, 59(6), 1210–1215. https://doi.org/10.1093/JAC/DKL509
  42. Zahn, M., Bhamidimarri, S. P., Baslé, A., Winterhalter, M., & van den Berg, B. (2016). Structural Insights into Outer Membrane Permeability of Acinetobacter baumannii. Structure (London, England: 1993), 24(2), 221–231. https://doi.org/10.1016/J.STR.2015.12.009
  43. Bhamidimarri, S. P., Zahn, M., Prajapati, J. D., Schleberger, C., Söderholm, S., Hoover, J., West, J., Kleinekathöfer, U., Bumann, D., Winterhalter, M., & van den Berg, B. (2019). A Multidisciplinary Approach toward Identification of Antibiotic Scaffolds for Acinetobacter baumannii. Structure (London, England: 1993), 27(2), 268-280.e6. https://doi.org/10.1016/J.STR.2018.10.021
  44. Borneleit, P., & Kleber, H.-P. (1991). The Outer Membrane of: Structure-Function Relationships. The Biology of Acinetobacter, 259–271. https://doi.org/10.1007/978-1-4899-3553-3_18
  45. Zahn, M., Bhamidimarri, S. P., Baslé, A., Winterhalter, M., & van den Berg, B. (2016). Structural Insights into Outer Membrane Permeability of Acinetobacter baumannii. Structure (London, England: 1993), 24(2), 221–231. https://doi.org/10.1016/J.STR.2015.12.009