STRUCTURAL CHARACTERIZATION OF SP1-DNA COMPLEXES
SP1-DNA 复合物的结构表征
基本信息
- 批准号:3307848
- 负责人:
- 金额:$ 13.08万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:1992
- 资助国家:美国
- 起止时间:1992-09-30 至 1996-08-31
- 项目状态:已结题
- 来源:
- 关键词:DNA DNA binding protein DNA footprinting biophysics calorimetry chemical binding circular dichroism gel mobility shift assay genetic promoter element intermolecular interaction nuclear magnetic resonance spectroscopy nucleic acid sequence polymerase chain reaction protein structure function solutions thermodynamics transcription factor
项目摘要
Synthesis of messenger RNA by RNA polymerase II requires the interaction
of a large array of auxiliary transcription factors that recognize and
bind to specific promoter DNA sequences located upstream of eukaryotic
genes. These transcription factors regulate the initiation of
transcription in a temporally ordered manner by assembling and engaging
the active transcription complex. In order to understand the detailed
roles played by transcription factors, efforts have been made to
fractionate the factors necessary to reconstitute transcriptional
activity in vitro. These experiments have resulted in the
identification, purification and characterization of one such
promoter-specific transcription factor, Sp l, from HeLa cells. Sp l
enhances transcription from a variety of viral and cellular genes by
binding to one or more "GC box" recognition elements (containing a
hexanuclear core GGGCGG) within the 5' flanking promoter sequences
through the use of three "zinc-finger" domains.
In general, DNA binding surfaces are designed to have a highly defined
preference for their cognate DNA binding site; Sp l is unique among
transcription factors identified to date in that it recognizes a host of
transcription activating binding sites that can be classified as either
high, medium or low affinity. We are therefore quite interested in
examining the binding of Sp1 to "GC box" sequences in order to define
those factors responsible for this unusually promiscuous sequence
recognition ability. The Spl system is amenable to detailed examination
owing to localization of its DNA binding properties to the "zinc-finger"
domain. This proposal describes an approach that will enable us to
exploit the relatively compact "zinc-finger" motif to define those
structural factors responsible for the recognition diversity of Sp 1.
Our objectives are: ( 1) to express short soluble Sp 1 fragments that
contain the three "zinc finger" domains, (2) to test the ability of these
fragments to duplicate the natural binding properties of intact Spl by
band shift assays, (3) to express and purify large quantities of those
peptides that exhibit Spl binding ability, (4) to quantitatively
characterize the interactions of these fragments with duplex
oligonucleotides by molecular biological (band shift assays, competition
assays, mutagenesis studies) and biophysical techniques (calorimetry,
NMR) in order to define the chemical/structural basis for the observed
diversity in DNA binding, and (5) to determine the solution structures of
the active Spl fragments by NMR spectroscopic techniques.
Our long term objective is to characterize the solution structures of a
number of Spl -DNA complexes using a series of high and medium affinity
DNA recognition sequences. Such data will not only contribute to our
understanding at the molecular level of how "zinc-finger" domains are
used in molecular recognition but also how this major structural motif is
able to recognize a highly variable set of DNA sequences.
RNA聚合酶II合成信使RNA需要相互作用
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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John P. Caradonna其他文献
Structure/function relationships in mononuclear non-heme iron enzymes
- DOI:
10.1016/0162-0134(95)97462-y - 发表时间:
1995-08-01 - 期刊:
- 影响因子:
- 作者:
Jeffrey M. Zaleski;Kelly E. Loeb;Allen M. Orville;T.J. Kappock;John P. Caradonna;John D. Lipscomb;Edward I. Solomon - 通讯作者:
Edward I. Solomon
John P. Caradonna的其他文献
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{{ truncateString('John P. Caradonna', 18)}}的其他基金
BIOLOGICAL OXIDATION OF PHENYLALANINE BY NONHEME IRON
非血红素铁对苯丙氨酸的生物氧化
- 批准号:
6090321 - 财政年份:2000
- 资助金额:
$ 13.08万 - 项目类别:
BIOLOGICAL OXIDATION OF PHENYLALANINE BY NONHEME IRON
非血红素铁对苯丙氨酸的生物氧化
- 批准号:
6520232 - 财政年份:2000
- 资助金额:
$ 13.08万 - 项目类别:
BIOLOGICAL OXIDATION OF PHENYLALANINE BY NONHEME IRON
非血红素铁对苯丙氨酸的生物氧化
- 批准号:
6636443 - 财政年份:2000
- 资助金额:
$ 13.08万 - 项目类别:
BIOLOGICAL OXIDATION OF PHENYLALANINE BY NONHEME IRON
非血红素铁对苯丙氨酸的生物氧化
- 批准号:
6363349 - 财政年份:2000
- 资助金额:
$ 13.08万 - 项目类别:
STRUCTURAL CHARACTERIZATION OF SP1-DNA COMPLEXES
SP1-DNA 复合物的结构表征
- 批准号:
2185857 - 财政年份:1992
- 资助金额:
$ 13.08万 - 项目类别:
STRUCTURAL CHARACTERIZATION OF SP1-DNA COMPLEXES
SP1-DNA 复合物的结构表征
- 批准号:
2185858 - 财政年份:1992
- 资助金额:
$ 13.08万 - 项目类别:
STRUCTURAL CHARACTERIZATION OF SP1-DNA COMPLEXES
SP1-DNA 复合物的结构表征
- 批准号:
3307847 - 财政年份:1992
- 资助金额:
$ 13.08万 - 项目类别:
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