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

Year 2023, Volume: 6 Issue: 2, 1529 - 1542, 05.07.2023

Özge Kaygusuz İzgördü , Cihan Darcan , Fatma Doğan Guzel

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.

Keywords

E. coli, A. baumannii, Ni-NTA, Protein Purification, OccAB1

Supporting Institution

TUBITAK

Project Number

117S114

Thanks

This work was supported by the TUBITAK project numbered 117S114. We thank TUBITAK for this support. The authors would like also to thank Araz N. Dizaji and Ikbal Agah İnce for helpful discussions during the preliminary studies.

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

Abstract

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.

Keywords

E. coli, A. baumannii, Ni-NTA, Protein saflaştırma, OccAB1

Project Number

117S114

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