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Title: DNA and Protein Footprinting Analysis of the Modulation of DNA Binding by the N-Terminal Domain of the Saccharomyces cervisiae TATA Binding Protein

Abstract

Recombinant full-length Saccharomyces cerevisiae TATA binding protein (TBP) and its isolated C-terminal conserved core domain (TBPc) were prepared with measured high specific DNA-binding activities. Direct, quantitative comparison of TATA box binding by TBP and TBPc reveals greater affinity by TBPc for either of two high-affinity sequences at several different experimental conditions. TBPc associates more rapidly than TBP to TATA box bearing DNA and dissociates more slowly. The structural origins of the thermodynamic and kinetic effects of the N-terminal domain on DNA binding by TBP were explored in comparative studies of TBPc and TBP by 'protein footprinting' with hydroxyl radical ({center_dot}OH) side chain oxidation. Some residues within TBPc and the C-terminal domain of TBP are comparably protected by DNA, consistent with solvent accessibility changes calculated from core domain crystal structures. In contrast, the reactivity of some residues located on the top surface and the DNA-binding saddle of the C-terminal domain differs between TBP and TBPc in both the presence and absence of bound DNA; these results are not predicted from the crystal structures. A strikingly different pattern of side chain oxidation is observed for TBP when a nonionic detergent is present. Taken together, these results are consistent with the N-terminal domainmore » actively modulating TATA box binding by TBP and nonionic detergent modulating the interdomain interaction.« less

Authors:
; ; ; ; ; ; ;
Publication Date:
Research Org.:
Brookhaven National Laboratory (BNL) National Synchrotron Light Source
Sponsoring Org.:
Doe - Office Of Science
OSTI Identifier:
929940
Report Number(s):
BNL-80535-2008-JA
Journal ID: ISSN 0006-2960; TRN: US200822%%1109
DOE Contract Number:  
DE-AC02-98CH10886
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemistry; Journal Volume: 46
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; AFFINITY; BEARINGS; CRYSTAL STRUCTURE; DETERGENTS; DNA; HYDROXYL RADICALS; KINETICS; MODULATION; OXIDATION; PROTEINS; REACTIVITY; RESIDUES; SACCHAROMYCES; SACCHAROMYCES CEREVISIAE; SOLVENTS; SURFACES; TBP; THERMODYNAMICS; TRIOCTYLPHOSPHINE SULFIDE; national synchrotron light source

Citation Formats

Gupta,S., Cheng, H., Mollah, A., Jamison, E., Morris, S., Chance, M., Khrapunov, S., and Brenowitz, M. DNA and Protein Footprinting Analysis of the Modulation of DNA Binding by the N-Terminal Domain of the Saccharomyces cervisiae TATA Binding Protein. United States: N. p., 2007. Web. doi:10.1021/bi7003608.
Gupta,S., Cheng, H., Mollah, A., Jamison, E., Morris, S., Chance, M., Khrapunov, S., & Brenowitz, M. DNA and Protein Footprinting Analysis of the Modulation of DNA Binding by the N-Terminal Domain of the Saccharomyces cervisiae TATA Binding Protein. United States. doi:10.1021/bi7003608.
Gupta,S., Cheng, H., Mollah, A., Jamison, E., Morris, S., Chance, M., Khrapunov, S., and Brenowitz, M. Mon . "DNA and Protein Footprinting Analysis of the Modulation of DNA Binding by the N-Terminal Domain of the Saccharomyces cervisiae TATA Binding Protein". United States. doi:10.1021/bi7003608.
@article{osti_929940,
title = {DNA and Protein Footprinting Analysis of the Modulation of DNA Binding by the N-Terminal Domain of the Saccharomyces cervisiae TATA Binding Protein},
author = {Gupta,S. and Cheng, H. and Mollah, A. and Jamison, E. and Morris, S. and Chance, M. and Khrapunov, S. and Brenowitz, M.},
abstractNote = {Recombinant full-length Saccharomyces cerevisiae TATA binding protein (TBP) and its isolated C-terminal conserved core domain (TBPc) were prepared with measured high specific DNA-binding activities. Direct, quantitative comparison of TATA box binding by TBP and TBPc reveals greater affinity by TBPc for either of two high-affinity sequences at several different experimental conditions. TBPc associates more rapidly than TBP to TATA box bearing DNA and dissociates more slowly. The structural origins of the thermodynamic and kinetic effects of the N-terminal domain on DNA binding by TBP were explored in comparative studies of TBPc and TBP by 'protein footprinting' with hydroxyl radical ({center_dot}OH) side chain oxidation. Some residues within TBPc and the C-terminal domain of TBP are comparably protected by DNA, consistent with solvent accessibility changes calculated from core domain crystal structures. In contrast, the reactivity of some residues located on the top surface and the DNA-binding saddle of the C-terminal domain differs between TBP and TBPc in both the presence and absence of bound DNA; these results are not predicted from the crystal structures. A strikingly different pattern of side chain oxidation is observed for TBP when a nonionic detergent is present. Taken together, these results are consistent with the N-terminal domain actively modulating TATA box binding by TBP and nonionic detergent modulating the interdomain interaction.},
doi = {10.1021/bi7003608},
journal = {Biochemistry},
number = ,
volume = 46,
place = {United States},
year = {Mon Jan 01 00:00:00 EST 2007},
month = {Mon Jan 01 00:00:00 EST 2007}
}