Related Experiment Videos
Autogenous translational operator recognized by bacteriophage T4 DNA polymerase
1Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder 80309.
Journal of Molecular Biology
|June 20, 1990
Summary
Bacteriophage T4 DNA polymerase (gp43) autoregulates its synthesis by binding to its mRNA, blocking translation. This autoregulation is separate from its replication function, suggesting a model linking synthesis control to demand.
Area of Science:
- Molecular Biology
- Virology
- Genetics
Background:
- Bacteriophage T4 DNA polymerase (gp43) synthesis is autogenously regulated.
- gp43 binds to its mRNA operator sequence, inhibiting translation.
- This autoregulation mechanism involves a specific mRNA hairpin-loop structure.
Purpose of the Study:
- To determine the binding affinity (Kd) of gp43 to its mRNA operator.
- To characterize the minimal operator sequence required for gp43 binding.
- To investigate the relationship between gp43 autoregulation and its replicative function.
Main Methods:
- Determined the dissociation constant (Kd) of the gp43-operator interaction.
- Identified the minimal 36-nucleotide operator sequence, including a hairpin and Shine-Dalgarno sequence.
- Constructed phage operator mutants to study in vivo effects.
Main Results:
- The Kd of the gp43-operator interaction was found to be 1.0 x 10(-9) M.
- The minimal operator sequence comprises a hairpin with a 5 base-pair helix and 8-nucleotide loop.
- Mutants overproducing gp43 showed no change in replication rates or phage yield.
- Replicative and autoregulatory functions were found to be mutually exclusive.
Conclusions:
- gp43 autoregulation is mediated by binding to a conserved hairpin-loop structure in its mRNA.
- Autoregulation and replication are distinct, mutually exclusive activities of the DNA polymerase.
- A model is proposed where gp43 synthesis is linked to replicative demand via autoregulation.