Research in my laboratory is focused in several broad areas: antigenic characterization and immune responses against pathogenic microorganisms and the application of such information to protective immunity and diagnostics; protein translocation and membrane biogenesis as related to bacterial stress tolerance; and studies of intra- and inter-molecular protein-protein interactions relevant for bacterial protein stability, function and adhesive interactions with host constituents.
The ribonucleotide-protein complex signal recognition particle (SRP) is conserved in all three domains of life and delivers membrane and secretory proteins to the cyoplasmic membrane or endoplasmic reticulum. Our laboratory was the first to demonstrate that this pathway is not essential for viability in a living organism and that the cariogenic bacterium Streptococcus mutans survives albeit with increased vulnerability to environmental stressors in the absence of the universally conserved minimal elements of this co-translational translocation pathway. Streptococci also encode two homologs of the evolutionary conserved YidC/Oxa/Alb family of membrane-associated chaperones expressed by bacteria, mitochonidria, and chloroplasts that mediate and contribute to the insertion of membrane proteins and multi subunit complexes, including components of the F1F0 ATP synthase. The more widely studied model organism Escherichia coli expresses a single YidC that like the SRP pathway is also necessary for viability in this species. Our results suggest functional overlap between the SRP pathway and YidC2 in S. mutans. We are currently exploring the roles of the SRP pathway and YidC homologs in membrane biogenesis and adaptation to environmental stress and evaluating the compensatory mechanisms by which streptococci can survive without molecules considered essential in other organisms.
Our laboratory is also defining chaperone interactions necessary for stability, maturation and surface expression of streptococcal surface molecules translocated by the post-translational general secretion pathway. We have characterized multiple conformational epitopes recognized by monoclonal antibodies against the P1 adhesin of S. mutans and demonstrated that the an intra-molecular interaction between the amino-terminal alanine-rich repeat and central proline-rich repeat domains is necessary for the structural integrity, surface localization and function of the molecule. Conserved architectural features among numerous gram-positive surface proteins indicate our system is a useful model to dissect the intracellular chaperone machinery involved in protein folding and trafficking. A gene discovery approach is being used to identify global changes in protein expression that occur when adhesin P1 is engaged with a physiological binding partner. Also, in collaboration with Dr. Champion Deivanayagam of the Center for Biophysical Sciences and Engineering at the University of Alabama we are utilizing X-ray crystallography in conjunction with protein interaction methodologies to define the structure of P1 and to identify the binding motif responsible for its interaction with the high molecular weight gp340 salivary agglutinin ligand.
In addition, our group has also used our panel of well characterized monoclonal antibodies against S. mutans P1, a recognized target of protective immunity and widely studied candidate vaccine antigen, to demonstrate that exogenously administered antibodies included as part of an immune complex prior to immunization can be used in a directed and practical manner to alter the host antibody response against the bacteria towards one of increased efficacy. The altered response mediated by several of the immunomodulatory antibodies demonstrates significantly improved inhibition of bacterial adherence. Work is underway to elucidate correlates of protection in vivo and to evaluate molecular mechanisms of immunomodulation, including induced changes in antigen conformation and exposure of cryptic epitopes and subsequent alterations in antigen processing and/or presentation. As it is becoming increasingly recognized that passively administered antibodies influence subsequent humoral as well as cell-mediated adaptive immune responses against the antigens to which they bind, this information is broadly relevant to active and passive vaccine approaches against both extracellular and intracellular pathogenic microorganisms.
Finally, we have demonstrated novel cell surface antigens associated with a highly pathogenic lineage of the neonatal pathogen Streptococcus agalactiae and identified their underlying genetic basis. Work is directed at understanding differences within this family of streptococcal and staphylococcal serine-rich repeat molecules with regard to pathogenesis and to determine potential utility as vaccine components or diagnostic markers. Previously, a domain necessary, but not sufficient for, human IgA Fc binding activity of the group B streptococcal beta antigen was identified and enabled a deletion construct to be engineered for use as a candidate immunogen and carrier in a multi-component vaccine preparation. My laboratory also collaborates with investigators who are using the immune response against microorganisms as a reporter system for the identification of genes expressed during in vivo infections.
- Larson, M., Rajashankar, K., Patel, M., Robinette, R., Crowley, P., Michalek, S., Brady, L.J., Deivanayagam, C. (2010) Elongated fimbiral structure of a streptococcal adhesin assembled by the high affinity association of alpha- and PPII-helices. Proc. Natl. Acad. Sci. March 15 (e-Pub ahead of print).
- Tavares, M.B., Silva, B.M., Calvacante, C.M., Souza, R.D., Luiz, W.B., Paccez, J.D., Crowley, P.J., Brady, L.J., Ferreira, L.C.S., and Ferreira, R.C.C. (2010) Induction of neutralizing antibodies in mice immunized with an amino-terminal polypeptide of Streptococcus mutans P1 protein produced by a recombinant Bacillus subtilis strain. FEMS Immunol. Med. Microbiol. In press.
- Robinette, R.R., Oli, M.W., McArthur, W.P., and Brady, L.J. (2009) Beneficial immunomodulation by Streptococcus mutans anti-P1 monoclonal antibodies is Fc-independent and correlates with increased exposure of a relevant target epitope. J. Immunol. 183: 4628-4638.
- Funes, S. Hasona, A., Grubbauer, C., Bauerschmitt, H., Kauff, F., Collins, R., Crowley, P.J., Palmer, S.R., Brady, L.J.* and Herrmann, J.M.* (*co-corresponding authors) (2009) Independent gene duplications of the YidC/Oxa/Alb3 family enabled a specialized co-translational function. Proc. Natl. Acad. Sci. 106(16): 6656-61.
- Crowley, P.C., Seifert, T.B., Isoda, R., van Tilburg, M., Oli, M.W., Robinette, R.A., McArthur, W.P., Bleiweis, A.S., and Brady, L.J. (2008) Requirements for surface expression and function of adhesin P1 from Streptococcus mutans. Infect. Immun. 76 (6): 2456-2468.
- Dong, X., Palmer, S.R., Hasona, A., Nagamori, S., Kaback, H.R., Dalbey, R.E., and Brady, L.J. (2008) Functional overlap but lack of complete cross-complementation of Streptococcus mutans and Escherichia coli YidC homologs. J. Bacteriol. 190 (7): 2458-2469.
- Zuobi-Hasona, K. and Brady, L.J. (2008) Isolation and solubilization of cellular membrane proteins from bacteria? In, Methods in Molecular Biology Volume 425: 2D PAGE: Sample Preparation and fractionation, Volume 2 (A. Posch, ed.) Humana Press, Totowa, New Jersey, 287-293.
- Ahn, S.J., Ahn, S.J., Wen, Z.T., Brady, L.J., Burne, R.A. (2008) Characteristics of biofilm formation by Streptococcus mutans in the presence of saliva. Infect Immun. 76 (9): 4259-68.
- Nobbs, A.H., Vajna, R.M., Johnson, J.R. Zhang, Y., Erlandsen, S.L., Oli, M.W., Kreth, J., Brady, L.J., and Herzberg, M.C. (2007) Consequences of a sortase A mutation in Streptococcus gordonii. Microbiology. Dec: 153: (12):4088-4097.
- Isoda, R., Robinette, R., Pinder, T.L., McArthur, W.P., and Brady L.J. (2007) Basis of beneficial immunomodulation by monoclonal antibodies against Streptococcus mutans adhesin P1, FEMS, Immunol Med Microbiol. 51 (2007) 102–111.
- McArthur, William P., Rhodin, Nikki R., Seifert, T.B., Oli, M.W., Robinette, R., Demuth, D., and Brady L.J. (2007) Characterization of epitopes Recognized by Anti-P1 Streptococcus mutans Monoclonal Antibodies, FEMS Immunology and Medical Microbiology FEMS, Immunol Med Microbiol. 50(3): 342-53
- Hasona, A., Zuobi-Hasona, K., Crowley, P.J., Abranches, J., Ruelf, M.A., Bleiweis, A.S., and Brady, L.J. (2007) Membrane composition changes and physiological adaptation by Streptococcus mutans signal recognition particle (SRP) pathway mutants, J. Bacteriol. 189(4):1219-1230.
- Oli, M.W., McArthur, W.P., & Brady, L.J. (2006) A whole cell BIAcore assay to evaluate P1-mediated adherence of Streptococcus mutans to human salivary agglutinin and inhibition by specific antibodies, J Microbiol Methods, 65(3): 503-511.
- Seifert, K.N., Adderson, E.E., Bohnsack, J.F., and Brady, L.J. (2006) A unique serine-rich repeat protein (Srr-2) and novel surface antigen (epsilon) associated with a virulent lineage of serotype III Streptococcus agalactiae, Microbiology 152:1029-1040.
- Hasona, A., Crowley, P.J., Levesque, C.M., Mair, R.W., Cvitkovitch, D.G., Bleiweis, A.S., & Brady, L.J. (2005) Streptococcal viability and diminished stress tolerance in mutants lacking the signal recognition particle pathway or YidC2. Proc Natl Acad Sci 102(48): 17464-17471.
- Zuobi-Hasona, K., Crowley, P.J., Hasona, A., Bleiweis, A.S., & Brady, L.J. (2005) Solubilization of cellular membrane proteins from Streptococcus mutans for two-dimensional gel electrophoresis, Electrophoresis 26(6): 1200-5.
- Brady, L.J. (2005) Antibody-mediated immunomodulation: a strategy to improve host responses against microbial antigens. Infect Immun. 73(2): 671-8.
- Oli, M.W., Rhodin, N., McArthur, W.P., & Brady, L.J. (2004) Re-directing the humoral immune response against Streptococcus mutans antigen P1 with monoclonal antibodies. Infect Immun, 72(12): 6951-60.
- Seifert, T.B., Bleiweis, A.S., & Brady, L.J. (2004) Contribution of the alanine-rich region of Streptococcus mutans P1 to antigenicity, surface expression, and interaction with the proline-rich repeat domain. Infect Immun, 72(8): 4699-706.
- Rhodin, N.R., Cutalo, J.M., Tomer, K.B., McArthur, W.P., & Brady, L.J. (2004) Characterization of the Streptococcus mutans P1 epitope recognized by immunomodulatory monoclonal antibody 6-11A. Infect Immun, 72(8): 4680-8.
- Crowley, P.J., Svensater, G., Snoep, J.L., Bleiweis, A.S., & Brady, L.J. (2004) An ffh mutant of Streptococcus mutans is viable and able to physiologically adapt to low pH in continuous culture. FEMS Microbiol Lett, 234(2): 315-24.
- Rhodin, N.R., Van Tilburg, M.L., Oli, M.W., McArthur, W.P., & Brady, L.J. (2004) Further characterization of immunomodulation by a monoclonal antibody against Streptococcus mutans antigen P1. Infect Immun, 72(1): 13-21.