Linked Life Data

2007130

Source:http://linkedlifedata.com/resource/pubmed/id/2007130

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
11
pubmed:dateCreated
1991-4-30
pubmed:abstractText
Native EcoRI DNA methyltransferase (Mtase, Mr 38,050) is proteolyzed by trypsin to generate an intermediate 36-kDa fragment (p36) followed by the formation of two polypeptides of Mr 23,000 and 13,000 (p23 and p13, respectively). Protein sequence analysis of the tryptic fragments indicates that p36 results from removal of the first 14 or 16 amino acids, p23 spans residues 15-216, and p13 spans residues 217-325. The relative resistance to further degradation of p23 and p13 suggests stable domain structures. This is further supported by the generation of similar fragments with SV8 endoprotease which has entirely different peptide specificities. Our results suggest the Mtase is a two-domain protein connected by a highly flexible interdomain hinge. The putative hinge region encompasses previously identified peptides implicated in AdoMet binding [Reich, N.O., & Everett, E. (1990) J. Biol. Chem. 265, 8929-8934] and catalysis [Everett et al. (1990) J. Biol. Chem. 265, 17713-17719]. Protection studies with DNA, S-adenosylmethionine (AdoMet), S-adenosylhomocysteine (AdoHcy), and sinefungin (AdoMet analogue) show that the Mtase undergoes significant conformational changes upon ligand binding. Trypsinolysis of the AdoMet-bound form of the Mtase generates different fragments, and the AdoMet-bound form is over 800 times more stable than unbound Mtase. The sequence-specific ternary complex (Mtase-DNA-sinefungin) is 2000 times more resistant to degradation by trypsin; cleavage eventually generates 26- and 12-kDa fragments which span residues 104-325 and 1-103, respectively (p26 and p12). The first 14 or 16 amino acids of the Mtase are not essential since p36 retains activity. Activity analysis of the p26 and p12 mixture also indicates retention of activity.(ABSTRACT TRUNCATED AT 250 WORDS)
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
19
pubmed:volume
30
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
2940-6
pubmed:dateRevised
2000-12-18
pubmed:meshHeading
pubmed:year
1991
pubmed:articleTitle
Structural and functional analysis of EcoRI DNA methyltransferase by proteolysis.
pubmed:affiliation
Department of Chemistry, University of California, Santa Barbara 93106.
pubmed:publicationType
Journal Article