HSD restriction-modification proteins partake in latent anticodon nuclease

Area of Science:

• Microbiology
• Molecular Biology
• Enzymology

Background:

• Phage T4 infection involves an anticodon nuclease (ACNase) that modifies host tRNA.
• The ACNase enzyme is encoded by the Escherichia coli prr locus in a latent form, activated by phage Stp protein.
• Latency is due to masking of the core PrrC factor by elements homologous to type I restriction-modification (Hsd) genes.

Purpose of the Study:

• To elucidate the mechanism of anticodon nuclease activation and its interaction with host factors.
• To investigate the role of E. coli Hsd proteins in the latency and activation of the anticodon nuclease.
• To explore the potential coupling of tRNA and DNA restriction systems.

Main Methods:

• Enzyme assays using cell extracts and purified components.
• Immunoassays employing Hsd antibodies.
• Genetic analysis of the prr locus and phage stp gene.

Main Results:

• Activation of latent ACNase in vitro required Stp, ATP, GTP, and endogenous DNA.
• A heat-stable E. coli factor could substitute for Stp, suggesting broader activation possibilities.
• Hsd antibodies recognized the holoenzyme but not the core PrrC, confirming Hsd's role in masking PrrC.

Conclusions:

• E. coli Hsd proteins are integral components of the latent ACNase complex, masking the PrrC core.
• Phage Stp protein, potentially with Hsd ligands, disrupts this masking interaction to activate the nuclease.
• The Hsd-PrrC interaction suggests a coordinated mechanism between prokaryotic DNA and tRNA restriction systems.