TAILIEUCHUNG - Substitution of the conserved cysteine with glycine (Cys82Gly) of Agmenellum quadruplicatum methylase Aqul (M. Aqul) is not cytotoxic to E. coli

Cytosine 5 DNA methyltransferases share ten conserved motifs. Motif IV contains an absolutely conserved proline-cysteine dipeptide. The cysteine residue of this motif is involved in catalysis by forming a covalent bond with the 6-position of cytosine prior to methyl group transfer. | Turk J Biol 25 (2001) 177-184 © TÜB‹TAK Substitution of the Conserved Cysteine With Glycine (Cys82Gly) of Agmenellum quadruplicatum Methylase Aqul (M. Aqul) is not Cytotoxic to E. coli Hatice PINARBAfiI Cumhuriyet University, Faculty of Medicine, Department of Biochemistry, 58140, Sivas-TURKEY Ergün PINARBAfiI Cumhuriyet University, Faculty of Medicine, Department of Medical Biology and Genetics, 58140, Sivas-TURKEY David P. HORNBY Krebs Institute for Biomolecular Research, Department of Molecular Biology, Universiyt of Sheffield, Western Bank, Sheffield. S10 2TN, UK Received: Abstract: Cytosine 5 DNA methyltransferases share ten conserved motifs. Motif IV contains an absolutely conserved proline-cysteine dipeptide. The cysteine residue of this motif is involved in catalysis by forming a covalent bond with the 6-position of cytosine prior to methyl group transfer. AquI DNA methyltransferase recognising the sequence CCCGGG is a heterodimer unlike other C5 DNA methyltransferases. We changed the conserved cysteine (Cys82) of to serine and glycine. The presence of mutations was confirmed by automated DNA sequencing. Mutants were tested by in vivo plasmid protection assay and also by transformation into mcrA+BC+ strain of E. coli. Replacement of the conserved cysteine with serine led to an apparent loss of the methyltransferring ability of the enzyme. Interestingly, it was found that substitution of cysteine with glycine is not cytotoxic to E. coli in the case of . Key Words: , mcr, DNA Methyltransferase, site-directed mutagenesis Agmenellum quadruplicatum Metiltransferaz› AquI () Korunmufl Sisteininin Glisinle De¤ifltirilmesi (Cys82Gly) E. coli ‹çin Sitotoksik De¤ildir Özet: Sitozin 5 DNA metiltransferazlar evrim süreci içinde korunmufl on motif içerirler. Motif IV kesinlikle korunmufl Prolin-Sistein dipeptid içerir. Bu motifdeki sistein amino asidi metil grup transferi için sitozinin 6. karbonu ile kovalent ba¤ oluflturarak .

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