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Utilizing steady‐state and time‐resolved fluorescence to probe DNA/Gata binding interactions.
Author(s) -
Lewis Jacquelyn,
Hartsell Lydia,
Kuiper Nora,
Krueger Brent,
Pikaart Michael
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.a1030-c
Subject(s) - dna , fluorescence anisotropy , chemistry , zinc finger , fluorescence , transcription factor , biophysics , steady state (chemistry) , microbiology and biotechnology , biochemistry , biology , gene , physics , quantum mechanics , membrane
The recruitment of transcription factor proteins is an essential step in the regulation of many processes including hematopoiesis—the production of red blood cells. One critical transcription factor in this process, GATA‐1, binds to the DNA sequence (A/T)GATA(A/G) via a zinc finger. The specificity of this interaction is governed by several key amino‐acid/nucleotide contacts, which were identified using gel electrophoresis (Mott, Bassman, and Pikaart, BBRC 316 (2004) –917). These studies are being extended using steady‐state and time‐resolved fluorescence anisotropy to characterize the thermodynamics of the GATA‐1/DNA interaction in a solution‐phase environment. The anisotropic properties of the fluorescent probe Pacific Blue (PB), both free in solution and conjugated to double‐stranded DNA, have been determined. Results thus far have shown sensitivity of PB to changing environmental factors, such as temperature. Anisotropy measurements of the GATA‐1/DNA‐PB complex are ongoing.