Structure of a staphylococcal phage connector protein

Barbora Břenková¹, Marta Šiborová¹, Jiří Nováček¹, Pavol Bárdy², Roman Pantůček², Pavel Plevka¹

¹Central European Institute of Technology, Masaryk University, Kamenice 753/5, 625 00 Brno, Czech Republic;

²Department of Experimental Biology, Faculty of Science, Masaryk University, Kotlářská 267/2, 611 37 Brno, Czech Republic; correspondence should be addressed to pavel.plevka@ceitec.muni.cz

Antibiotic resistance of microorganisms is a major threat accounting for a rising number of deaths [1]. Bacteriophages offer an alternative to conventional antimicrobial treatment. Phage phi812 is a lytic virus from the Myoviridae family infecting up to 95 % of Staphylococcus aureus strains, including MRSA [2] and its structure was previously determined by cryo-electron microscopy [3]. Here we present our work towards resolving the structure of protein gp99 forming the connector region. The protein may play a role in genome release regulation during infection. Gp99 was overexpressed in E. coli and purified by affinity and size exclusion chromatography. A suitable crystallization condition was found and X-ray diffraction data with a resolution of 2.2 Å were collected. To solve the phase problem, a variety of methods had been tried, including molecular replacement, heavy atom soaking, co-crystallization, and seleno-methionine incorporation. The structure of gp99 will help explain the mechanism of bacterial infection by bacteriophage.

1. “Global action plan on antimicrobial resistance.” Geneva: World Health Organization; 2015, https://www.who.int/antimicrobial-resistance/publications/global-action-plan/en/.

2. Pantůček, R., Rosypalová, A., Doškař, J., Kailerová, J., Růžičková, V., Borecká, P., Snopková, S., Horváth, R., Götz, F., Rosypal, S. “The Polyvalent Staphylococcal Phage Phi 812: Its Host-Range Mutants and Related Phages.” Virology., U.S. National Library of Medicine, 5 July 1998, www.ncbi.nlm.nih.gov/pubmed/9657943.

3. Nováček, J., Šiborová, M., Benešík, M., Pantůček, R., Doškař, J., Plevka, P. “Structure and Genome Release of Twort-like Myoviridae Phage with a Double-Layered Baseplate.” Proceedings of the National Academy of Sciences of the United States of America., U.S. National Library of Medicine, 16 Aug. 2016, www.ncbi.nlm.nih.gov/pubmed/27469164.