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oai:www.ideals.illinois.edu:2142/85456 2023-07-11 col_2142_5131 col_2142_14828 com_2142_5130 com_2142_14827 com_2142_14793 com_2142_8903

Ha, Taekjip McKinney, Mary Cathleen 2015-09-25T22:46:09Z 2015-09-25T22:46:09Z 10000-01-01 2006 2006 DNA replication is a complex process involving many proteins that must be performed for a cell to proliferate. In this work, we will study three members of the replication system: single-stranded DNA, single-stranded DNA binding proteins, and the sliding clamp and clamp loader proteins. Fluorescence studies have been widely used to study the properties of biomolecules and here we will use both Fluorescence Polarization Anisotropy (FPA) and Fluorescence Resonance Energy Transfer (FRET) to study these biomolecules. First, we present a method by which single molecule FRET measurements can be used to determine the persistence length of single-stranded DNA. Our results indicate that the persistence length decreases from 3.0nm to 1.5nm as the salt concentration increases from 50mM NaCl to 2M NaCl. Furthermore, we use fluoresce anisotropy and FRET measurements in ensemble studies to examine properties of single stranded DNA binding proteins from archaeal organisms. Our results will show that the Archaea have exploited the single stranded DNA binding fold motif unlike the other domains of life to create a wide variety of proteins that have different binding properties leading to the probable conclusion that many of the proteins area a result of gene duplication and recombination events. Finally, we will examine the properties of the clamp loader protein as it loads a fluorescently labeled clamp to DNA on both the ensemble and single molecule measurements. Our findings show that loading the clamp is a multi-step process with different conformational states that can be observed for the first time at a single molecule level. We also will examine mutations of the clamp loader protein to bring further insight into the not fully understood Methanosarcina acetivorans clamp loader. Made available in DSpace on 2015-09-25T22:46:09Z (GMT). No. of bitstreams: 2 license.txt: 4848 bytes, checksum: 96035ab3f5e1c23cc7138a224ce498bd (MD5) 3242936.pdf: 3057992 bytes, checksum: ea876b5f959a0504040a17cfb841c4b3 (MD5) Previous issue date: 2006 Embargo set by: Seth Robbins for item 86737 Lift date: Forever Reason: Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs U of I Only 108 p. Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2006. http://hdl.handle.net/2142/85456 (MiAaPQ)AAI3242936 eng Biophysics, General Single -Molecule and Ensemble Fluorescence Studies of Single -Stranded DNA and Replication Proteins text Biophysics and Computational Biology Biophysics and Computational Biology University of Illinois at Urbana-Champaign Dissertation Ph.D.