The role of the preserved sequences of Dam methylase

Abstract

We have undertaken a site directed mutational analysis of two of the preserved regions in the amino acid sequence of Dam methylase in order to characterize their role. Mutations in region IV (sequence DPPY) abolish catalytic activity and greatly affect AdoMet crosslinking. Mutants in region III display a lowered specific activity with an unchanged AdoMet crosslinking capacity. We have also made a series of deletions both at the N and C terminal parts of the protein, which have been found to provide inactive enzyme. We discuss the significance of these results for the understanding of the functional properties of the enzyme.

Dam methylase fromEscherichia coli: kinetic studies using modified DNA oligomers: hemimethylated substrates

“…Biochemical studies on EcoRI Mtase (M-EcoRI) (9-13), M*EcaI (14,15), Dam Mtase (16)(17)(18)(19)(20) and M-MvaI (21) have been reported. A mechanism for methylation by M*EcoRI has been proposed (12,22,23).…”

Functional Roles of Conserved Amino Acid Residues in DNA Methyltransferases Investigated by Site-directed Mutagenesis of theEcoRV Adenine-N6-methyltransferase

“…Consequently, these additional motifs only recently could be identified in structure-guided alignments (21). Whereas some amino acid residues within the FXGXG and DPPY motifs have already been investigated by site-directed mutagenesis (22)(23)(24)(25), no mutational studies have been carried out so far to test the functional roles of conserved amino acid residues within the other motifs either in C-or N-MTases.…”

Dam methyltransferase from Escherichia coLi: sequence of a peptide segment involved in S-adenosyl-methionine binding

“…Site-directed mutagenesis experiments corroborated these findings: a truncated Dam methylase mutant shortened by ten N-terminal amino acids could not be expressed at all or only as insoluble inclusion bodies (20).…”