Identification of a highly conserved domain in the EcoRII methyltransferase which can be photolabeled with S-adenosyl-L-(methyl-3H)methionine. Evidence for UV-induced transmethylation of cysteine 186

Journal Article · Fri Feb 15 04:00:00 EST 1991 · Journal of Biological Chemistry; (USA)

OSTI ID:6056777

Som, S; Friedman, S ^[1]

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1. State Univ. of New York Health Science Center, Brooklyn (USA)

DNA methyltransferases can be photolabeled with S-adenosyl-L-methionine (AdoMet). Specific incorporation of radioactivity has been demonstrated after photolabeling with either (methyl-3H)AdoMet or (35S)AdoMet. The labeling is believed to occur at the AdoMet binding site. With the purpose of localizing the site responsible for (methyl-3H)AdoMet photolabeling, we cleaved the labeled EcoRII methyltransferase by chemical and enzymatic reactions and isolated the radiolabeled peptides by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high pressure liquid chromatography. The labeled peptides were identified by amino-terminal sequencing. A common region was localized which accounted for 65-70% of the total label. This region includes a highly conserved core sequence present in all DNA (cytosine 5)-methyltransferases. One such fragment was digested further with chymotrypsin, and amino acid analysis of the resulting 3H-labeled peptide was consistent with the sequence Ala-Gly-Phe-Pro-(Cys)-Gln-Pro-Phe-Ser-Leu. However, the cysteine residue was not recovered as carboxymethylcysteine. The Pro-Cys bond was found to be protected from cleavage at cysteine residues after cyanylation. These results suggest that the cysteine residue is modified by the labeling reaction. The chymotryptic fragment was hydrolyzed enzymatically to single amino acids, and the labeled amino acid was identified as S-methylcysteine by thin layer chromatography. These results indicate that the cysteine residue is located at or close to the AdoMet binding site of EcoRII methyltransferase.

• OSTI ID: 6056777
• Journal Information: Journal of Biological Chemistry; (USA), Journal Name: Journal of Biological Chemistry; (USA) Vol. 266:5; ISSN 0021-9258; ISSN JBCHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
560120 -- Radiation Effects on Biochemicals
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59 BASIC BIOLOGICAL SCIENCES
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
AMINO ACID SEQUENCE
AMINO ACIDS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CARBOXYLIC ACIDS
CHEMICAL REACTIONS
CHROMATOGRAPHY
CYSTEINE
DAYS LIVING RADIOISOTOPES
DNA
DRUGS
ELECTROMAGNETIC RADIATION
ELECTROPHORESIS
ENZYMES
EVEN-ODD NUCLEI
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPES
LABELLING
LIGHT NUCLEI
LIPOTROPIC FACTORS
LIQUID COLUMN CHROMATOGRAPHY
MEMBRANE PROTEINS
METHIONINE
METHYL TRANSFERASES
METHYLATION
MOLECULAR STRUCTURE
NUCLEI
NUCLEIC ACIDS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PROTEINS
RADIATIONS
RADIOISOTOPES
RECEPTORS
SEPARATION PROCESSES
SULFUR 35
SULFUR ISOTOPES
THIN-LAYER CHROMATOGRAPHY
THIOLS
TRACER TECHNIQUES
TRANSFERASES
TRITIUM COMPOUNDS
ULTRAVIOLET RADIATION