Linked Life Data

2793867

Source:http://linkedlifedata.com/resource/pubmed/id/2793867

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
29
pubmed:dateCreated
1989-11-17
pubmed:abstractText
Radiosequence analysis of peptide fragments of the estrogen receptor (ER) from MCF-7 human breast cancer cells has been used to identify cysteine 530 as the site of covalent attachment of an estrogenic affinity label, ketononestrol aziridine (KNA), and an antiestrogenic affinity label, tamoxifen aziridine (TAZ). ER from MCF-7 cells was covalently labeled with [3H]TAZ or [3H]KNA and purified to greater than 95% homogeneity by immunoadsorbent chromatography. Limit digest peptide fragments, generated by prolonged exposure of the labeled receptor to trypsin, cyanogen bromide, or Staphylococcus aureus V8 protease, were purified to homogeneity by high performance liquid chromatography (HPLC), and the position of the labeled residue was determined by sequential Edman degradation. With both aziridines, the labeled residue was at position 1 in the tryptic peptide, position 2 in the cyanogen bromide peptide, and position 7 in the V8 protease peptide. This localizes the site of labeling to a single cysteine at position 530 in the receptor sequence. The identity of cysteine as the site of labeling was confirmed by HPLC comparison of the TAZ-labeled amino acid (as the phenylthiohydantoin and phenylthiocarbamyl derivatives) and the KNA-labeled amino acid (as the phenylthiocarbamyl derivative) with authentic standards prepared by total synthesis. Cysteine 530 is located in the hormone binding domain of the receptor, near its carboxyl terminus. This location is consistent with earlier studies using sodium dodecyl sulfate-polyacrylamide gel electrophoresis to analyze the size of the proteolytic fragments containing the covalent labeling sites for TAZ and KNA and the antigen recognition sites for monoclonal antibodies. The fact that both the estrogenic and antiestrogenic affinity labeling agents react covalently with the same cysteine indicates that differences in receptor-agonist and receptor-antagonist complexes do not result in differential covalent labeling of amino acid residues in the hormone binding domain.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
http://linkedlifedata.com/resource/pubmed/chemical/Affinity Labels, http://linkedlifedata.com/resource/pubmed/chemical/Aziridines, http://linkedlifedata.com/resource/pubmed/chemical/Cyanogen Bromide, http://linkedlifedata.com/resource/pubmed/chemical/Cysteine, http://linkedlifedata.com/resource/pubmed/chemical/Ketones, http://linkedlifedata.com/resource/pubmed/chemical/Peptide Fragments, http://linkedlifedata.com/resource/pubmed/chemical/Receptors, Estrogen, http://linkedlifedata.com/resource/pubmed/chemical/Serine Endopeptidases, http://linkedlifedata.com/resource/pubmed/chemical/Tamoxifen, http://linkedlifedata.com/resource/pubmed/chemical/Trypsin, http://linkedlifedata.com/resource/pubmed/chemical/glutamyl endopeptidase, http://linkedlifedata.com/resource/pubmed/chemical/ketononestrol aziridine, http://linkedlifedata.com/resource/pubmed/chemical/tamoxifen aziridine
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
15
pubmed:volume
264
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
17476-85
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1989
pubmed:articleTitle
Identification of cysteine 530 as the covalent attachment site of an affinity-labeling estrogen (ketononestrol aziridine) and antiestrogen (tamoxifen aziridine) in the human estrogen receptor.
pubmed:affiliation
Department of Physiology and Biophysics, University of Illinois, Urbana 61801.
pubmed:publicationType
Journal Article, Comparative Study, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't