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Related Experiment Video

Updated: May 13, 2026

Sequence-specific Labeling of Nucleic Acids and Proteins with Methyltransferases and Cofactor Analogues

Sequence-specific Labeling of Nucleic Acids and Proteins with Methyltransferases and Cofactor Analogues

Published on: November 22, 2014

High frequency mutagenesis by a DNA methyltransferase.

J C Shen¹, W M Rideout, P A Jones

¹Department of Biochemistry and Molecular Biology, Kenneth Norris, Jr., Comprehensive Cancer Center, University of Southern California, School of Medicine, Los Angeles 90033.

|December 24, 1992

Summary

HpaII methylase induces C to U mutations in DNA. This enzyme

Area of Science:

Molecular Biology
Enzymology
Genetics

Background:

DNA methylation is crucial for gene regulation.
DNA (cytosine-5)-methyltransferases (DNMTs) are enzymes that catalyze DNA methylation.
HpaII methylase (M. HpaII) is a DNMT that recognizes the CCGG sequence.

Purpose of the Study:

To investigate the mutagenic potential of HpaII methylase.
To explore the mechanism of M. HpaII-induced mutations.
To determine the role of S-adenosylmethionine (SAM) in M. HpaII activity.

Main Methods:

Incubation of a mutant pSV2-neo plasmid with M. HpaII in vitro.
Transformation of the modified plasmid into bacteria lacking uracil-DNA glycosylase.
Mutation frequency analysis at varying SAM concentrations.

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Main Results:

M. HpaII directly induced a high frequency of C to U transition mutations in double-stranded DNA.
A 10^4-fold increase in reversion rate was observed in bacteria lacking uracil-DNA glycosylase.
Mutation frequency was highly sensitive to SAM concentration, decreasing at >300 nM.

Conclusions:

M. HpaII can act as a mutagen by inducing C to U transitions.
Enzymatic deamination by M. HpaII may contribute to mutational hotspots at CpG sites.
The findings support models of covalent complex formation between methyltransferases and cytosine.