John J. Dunn Member of the Brookhaven Council Biology Department, 463 Brookhaven National Laboratory Upton, NY 11973-5000 tel: (631) 344-3012 fax: (631) 344-3407 jdunn@bnl.gov The Group: John J. Dunn 334-3012 Barbara N. Lade 334-4690 Laura-Li Loffredo 334-4960 Beth Mulligan 334-4960 Dimitris Papamichail 334-4270 Judith A. Romeo 344-7430

Research Interests: Genome Signature Tags: It is well established that interactions between transcription factors and their cognate DNA binding sites form fundamental combinatorial networks within cells that control critical steps in gene expression. Currently, our understanding of how cells encode the diversity of information about where and when genes will be expressed is very limited. Cracking this 'regulatory code' by computational analysis and functional assays is a major problem in biology. The Dunn lab has combined Genomic Signature Tags (GSTs) technology, a method for identifying and quantitatively analyzing genomic DNAs originally devised in our lab, with chromatin immunoprecipitation (ChIP) assays to develop a high-throughput, direct, sequence-based approach at the whole genome level experimental data for delimiting the positions of bound regulatory proteins on prokaryotic or eukaryotic DNAs. In this protocol, concatemers of 20-21 bp long GST's, derived from ChIP precipitated DNA fragments, are sequenced en masse to identify the genomic segments originally present in these complexes. A long-range goal is to develop computational tools to model these protein-DNA interactions at the genome level and to make qualitative predictions on gene expression shifts that would be expected to emerge from specific pathway modulations. Affibody Technology: ChIP asays typically use polyclonal or monoclonal antibodies raised against a specific transcription factor or chromatin-associated protein. The step-by- step approach required to produce the antibodies is costly and inefficient. One current effort is to determine, in collaboration with Brian Kay, ANL, whether phage-displayed peptides and scFv antibodies (collectively termed affibodies) might replace antisera for ChIP experiments. Affibody libraries would be a much more economical and speedy method for obtaining ChIP fractions for additional experiments if this were the case. Zoonotic pathogens: The aim of this project is to modify our GST technology for profiling genomic DNAs to permit rapid, cultivation-free differential detection and identification of biothreat infectious agents in their natural environments, including intermediate infected hosts and clinical specimens from humans or infected animals. This project will test the utility of Pyrosequencing, a new DNA sequencing technique that is based on the detection of released pyrophosphate (PPi) during DNA synthesis, to rapidly identify zoonotic pathogens and biothreat agents such as foot-and-mouth disease virus. Lyme Disease: In an ongoing collaborative effort, we are conducting genomic and proteomic analyses of the Lyme disease spirochete, Borrelia burgdorferi to understand how specific gene products participate in immune evasion and pathogenicity. Our laboratory group has determined the structure of the outer surface protein A (OspA) of B. burgdorferi, the basis for the first FDA approved vaccine to protect humans against Lyme disease. We used this information to bioengineer chimer anti-OspA vaccine candidates for use in Eurasia where heterogeneity of the OspA lipoprotein limits the effectiveness of the first generation vaccine that was developed for use in the U.S.A. Recently, we also determined the structure of outer surface protein C, a potential second generation vaccine candidate, and showed that variations in the amino-acid sequence of OspC are correlated with differences in B. burgdorferi infectivity in humans. Currently, we are investigating whether complexes between OspC and ligands on the surface of eukaryotic cells are formed and whether they can help explain why only some types of OspC are found associated with invasive strains.

Becker M, Bunikis J, Lade BD, Dunn JJ, Barbour AG, and Lawson CL. Structural investigation of Borrelia burgdorferi OspB, a bactericidal Fab target. J Biol Chem. 280(17), 17363-17370 (2005). Medline Abstract PDB files: 1P4P, 1RJL;  -> Jmol viewer

Kumaran D, Eswaramoorthy S, Luft B, Koide S, Dunn JJ, Lawson CL, and Swaminathan S. Crystal structure of outer surface protein C (OspC) from the Lyme disease spirochete, Borrelia burgdorferi. EMBO J. 20, 971-978 (2001). Medline Abstract; PDB files: 1F1M, 1GGQ;  -> Jmol viewer

Ding W, Huang X, Yang X, Dunn JJ, Luft BJ, Koide S, and Lawson CL Structural identification of a key protective B-cell epitope in Lyme disease antigen OspA. J Mol Biol. 302, 1153-1164 (2000). Medline Abstract; PDB file 1FJ1;  -> Jmol viewer

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