caa a22 4500 60624252X CHVBK 20210128101319.0 cr unu---uuuuu 210128e20150901xx s 000 0 eng 10.1007/s12602-015-9192-8 doi (NATIONALLICENCE)springer-10.1007/s12602-015-9192-8 A Moonlighting Enolase from Lactobacillus gasseri does not Require Enzymatic Activity to Inhibit Neisseria gonorrhoeae Adherence to Epithelial Cells [Elektronische Daten] [Rachel Spurbeck, Paul Harris, Kannan Raghunathan, Dennis Arvidson, Cindy Arvidson] Enolases are generally thought of as cytoplasmic enzymes involved in glycolysis and gluconeogenesis. However, several bacteria have active forms of enolase associated with the cell surface and these proteins are utilized for functions other than central metabolism. Recently, a surface-associated protein produced by Lactobacillus gasseri ATCC 33323 with homology to enolase was found to inhibit the adherence of the sexually transmitted pathogen, Neisseria gonorrhoeae, to epithelial cells in culture. Here, we show that the protein is an active enolase in vitro. A recombinantly expressed, C-terminal His-tagged version of the protein, His6-Eno3, inhibited gonococcal adherence. Assays utilizing inhibitors of enolase enzymatic activity showed that this inhibitory activity required the substrate-binding site to be in an open conformation; however, the enolase enzymatic activity of the protein was not necessary for inhibition of gonococcal adherence. An L. gasseri strain carrying an insertional mutation in eno3 was viable, indicating that eno3 is not an essential gene in L. gasseri 33323. This observation, along with the results of the enzyme assays, is consistent with reports that this strain encodes more than one enolase. Here we show that the three L. gasseri genes annotated as encoding an enolase are expressed. The L. gasseri eno3 mutant exhibited reduced, but not abolished, inhibition of gonococcal adherence, which supports the hypothesis that L. gasseri inhibition of gonococcal adherence is a multifactorial process. Springer Science+Business Media New York, 2015 Neisseria gonorrhoeae nationallicence Hec-1-B cells nationallicence Sexually transmitted infection nationallicence Vaginal lactobacilli nationallicence Probiotics nationallicence Enolase nationallicence Spurbeck Rachel The Genetics Program, Michigan State University, East Lansing, MI, USA aut Harris Paul Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut Raghunathan Kannan Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut Arvidson Dennis Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut Arvidson Cindy Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut Probiotics and Antimicrobial Proteins Springer US; http://www.springer-ny.com 7/3(2015-09-01), 193-202 1867-1306 7:3<193 2015 7 12602 https://doi.org/10.1007/s12602-015-9192-8 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s12602-015-9192-8 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Spurbeck Rachel The Genetics Program, Michigan State University, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Harris Paul Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Raghunathan Kannan Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Arvidson Dennis Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Arvidson Cindy Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Rd., Biomedical Physical Sciences Rm 5192, 48824-1101, East Lansing, MI, USA aut NATIONALLICENCE 773 0- Probiotics and Antimicrobial Proteins Springer US; http://www.springer-ny.com 7/3(2015-09-01), 193-202 1867-1306 7:3<193 2015 7 12602