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Crystal structure of a replication fork single-stranded DNA binding protein (T4 gp32) complexed to DNA
Y Shamoo1, A M Friedman, M R Parsons
1Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520-8114, USA.
Nature
|July 27, 1995
Summary
The bacteriophage T4 gp32 protein binds single-stranded DNA (ssDNA) to ensure DNA replication, repair, and recombination. Its crystal structure reveals how gp32 acts as a chaperone, facilitating DNA strand movement.
Area of Science:
- Molecular Biology
- Structural Biology
- Virology
Background:
- The single-stranded DNA (ssDNA) binding protein gp32 is crucial for bacteriophage T4 DNA replication, recombination, and repair.
- In vivo, gp32 binds ssDNA at the replication fork, enhancing replisome processivity and accuracy significantly.
Purpose of the Study:
- To determine the crystal structure of the gp32 DNA binding domain complexed with ssDNA.
- To elucidate the molecular mechanisms by which gp32 interacts with and modulates ssDNA dynamics.
Main Methods:
- X-ray crystallography was used to determine the structure of the gp32 ssDNA complex at 2.2 Å resolution.
Main Results:
- The crystal structure revealed an ssDNA binding cleft formed by three subdomains, featuring a positively charged surface and hydrophobic pockets.
- Weak electron density for ssDNA suggests significant mobility, with the phosphate backbone interacting with the protein's electropositive cleft and bases contacting hydrophobic pockets.
Conclusions:
- Gp32 functions as a sequence-independent ssDNA chaperone, enabling the largely unstructured ssDNA to slide through its binding cleft.
- The structural data provides insights into gp32's roles in DNA metabolism, including template configuration, secondary structure melting, nuclease protection, and homologous recombination facilitation.