Linked Life Data

20715761

Source:http://linkedlifedata.com/resource/pubmed/id/20715761

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
35
pubmed:dateCreated
2010-8-31
pubmed:abstractText
X-ray structural studies revealed two conformations of the epidermal growth factor receptor (EGFR) ectodomain (ECD): a compact, tethered conformation in the absence of EGF and an untethered or extended conformation in the presence of EGF. An EGFR-ECD derivative with a monomeric red fluorescent protein (mRFP) at the N-terminus and an enhanced green fluorescent protein (eGFP) at the C-terminus (dual-tag-EGFR-ECD) was created and characterized. The dual-tag-EGFR-ECD construct was shown to have high affinity (nanomolar range) for both EGF and EGFR monoclonal antibody (mAb528). The dual-tag-EGFR-ECD was further characterized by fluorescence-detected analytical ultracentrifugation, lifetime FRET, and fluorescence anisotropy. We found no evidence of a tethered unliganded conformation, nor did we observe a large shape change upon ligand binding as predicted by the crystal models. Increases in steady-state anisotropy upon binding of EGF to the dual-tag-EGFR-ECD were observed and interpreted as changes in the protein flexibility and dynamics. We conclude the fluorescent protein tags perturb the EGFR-ECD structure, making it extended with a 50-fold higher affinity for EGF relative to that of the nontagged EGFR-ECD.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
1520-4995
pubmed:author
pubmed:issnType
Electronic
pubmed:day
7
pubmed:volume
49
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
7459-66
pubmed:meshHeading
pubmed:year
2010
pubmed:articleTitle
Creation and biophysical characterization of a high-affinity, monomeric EGF receptor ectodomain using fluorescent proteins.
pubmed:affiliation
Ludwig Institute for Cancer Research, P.O. Box 2008, Royal Melbourne Hospital, Victoria 3050, Australia.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't