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Specific interaction between a Saccharomyces cerevisiae protein and a DNA element associated with certain
S Eisenberg1, C Civalier, B K Tye
1Department of Microbiology, School of Medicine, University of Connecticut Health Center, Farmington 06032.
Summary
Researchers identified OBF1, a protein from yeast that binds to specific DNA sequences at autonomously replicating sequences (ARS). This protein, OBF1 (origin binding factor), may play a role in initiating DNA replication at certain ARS sites.
Area of Science:
- Molecular Biology
- Yeast Genetics
- DNA Replication
Background:
- Autonomously replicating sequences (ARS) are crucial for DNA replication initiation in yeast.
- Specific DNA-binding proteins are essential for regulating ARS function.
Purpose of the Study:
- To isolate and characterize a protein that binds to ARS120.
- To investigate the role of this protein in DNA replication.
Main Methods:
- Protein isolation and purification from Saccharomyces cerevisiae.
- DNase I footprinting to identify the DNA binding site.
- Affinity purification using a synthetic oligonucleotide repeat.
- Electrophoretic mobility shift assays (EMSAs) to confirm binding specificity.
Main Results:
- A novel protein, OBF1 (origin binding factor), was purified.
- OBF1 specifically binds to a 26-base-pair sequence within ARS120.
- OBF1 binds to a subset of ARS elements, but not ARS1 or mating type loci ARSs.
- The identified binding site is sufficient for OBF1 recognition.
Conclusions:
- OBF1 is a sequence-specific DNA-binding protein involved with ARS elements.
- OBF1 may function in the initiation of DNA replication at specific ARS loci in yeast.