Novel non-specific DNA adenine methyltransferases

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Bacteriophage Mu's mom gene encodes a protein that modifies adenine in viral DNA, protecting it from restriction endonucleases.
• Mu-like prophages in Haemophilus influenzae and Neisseria meningitidis lack the mom gene.

Purpose of the Study:

• To investigate the function of genes found in Mu-like prophages at the mom gene locus.
• To characterize the enzymatic activity and substrate specificity of the novel DNA adenine methyltransferases.

Main Methods:

• Gene identification and sequence analysis of Mu-like prophages.
• In vitro and in vivo DNA methylation assays.
• Kinetic analysis of oligonucleotide methylation.
• Restriction enzyme digestion assays on modified plasmid DNA.

Main Results:

• Mu-like prophages carry genes (hin1523, nma1821, hia5) encoding proteins homologous to DNA adenine N(6)-methyltransferases.
• These enzymes efficiently methylate adenine residues in DNA to N(6)-methyladenine, both in vitro and in vivo.
• Hia5 and Hin1523 exhibit broad substrate specificity, potentially methylating most adenine residues in DNA.
• Overexpression of these methyltransferases in E. coli resulted in plasmid DNA resistant to various restriction enzymes.

Conclusions:

• Novel DNA adenine methyltransferases in Mu-like prophages provide a defense mechanism against restriction enzymes, analogous to the mom gene.
• These enzymes represent a new class of DNA methyltransferases with broad substrate specificity.
• Understanding these enzymes could have implications for genetic engineering and microbial defense systems.