The aim of the study was to investigate quaternary structure of Omp85 outer membrane protein of Y. pestis, because of its potential use as a potential therapeutic target. Bacterial outer membrane proteins are suitable targets for the antimicrobial drugs and vaccine development. In Gram-negative bacteria, outer membrane protein, Omp85 is an integral part of the protein machinery. Inhibiting and disabling of Omp85 has a negative effect on the secretion of virulence factors, as well as a bactericidal effect. Previous research has identified components of the Omp85 complex and has started to yield structural insights into Omp85 family proteins, the quaternary structure of Omp85 itself remains under investigation. We present a quaternary structure study of Yersinia pestis Omp85 in vitro and in vivo using multiple approaches. Full-length Omp85, purified without denaturation, is shown to form monomers, dimers and tetramers in vitro using cross-linking and sedimentation studies. In vivo studies, using Western blot of outer membranes resolved on semi-native LDS-PAGE, indicate that Omp85 formed tetramers. Our studies contribute to a better understanding of Omp85 structure and mechanism. The applied relevance of this study has potential use in the development of new vaccines and antimicrobials, which may be potent weapons in the fight against bacterial infections.