SELENO-METHIONINE DERIVATIVE OF THE KH3 DOMAIN OF THE FUSE BINDING PROTEIN

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The FUSE Binding Protein (FBP) binds to the far upstream element (FUSE) of the c-myc oncogene, resulting in enhancement of c-myc transcription. This binding is mediated by four KH domains of FBP, each of which recognizes a single stranded consensus sequence within the FUSE. Understanding how KH domains recognize and discriminate amongst DNA species can provide insight into myc activation and processes resulting from it, such as cellular proliferation and tumorigenesis. NMR structures of the third and fourth KH domains (KH3 and KH4) of FBP, each bound to a cognate ssDNA sequence have been determined (Braddock et. al. Nature 415, pgs. 1051-1056, 2002); however, no crystallographic structure has been determined thus far. This work is part of ongoing efforts to design small molecules that inhibit DNA binding to FBP. The c-myc protein is important for cellular proliferation, but must be properly regulated to prevent excess growth and tumor formation. Because FBP-binding to the FUSE promotes c-myc transcription, disruption of this interaction in cases of c-myc over-activity may result in arresting, or the reversal of, tumor formation. This structure can also yield insight into the molecular recognition that is at the core of these important biological processes. It can also reveal the role played by water, which is not seen in the NMR structures, in this interaction. We have grown crystals of the KH3 domain bound to a SELEX-optimzed heptameric ssDNA and collected data at station X29 of the NSLS. A data set was processed to 2.6 Angstroms; however, attempts to solve the structure by molecular replacement failed. We propose to collect data from crystals of seleno-methionine KH3 domain bound to the same heptameric DNA sequence in order to solve the structure using selenium MAD.