Linked Life Data

8248098

Source:http://linkedlifedata.com/resource/pubmed/id/8248098

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
7
pubmed:dateCreated
1994-1-6
pubmed:abstractText
In order to probe the structural constraints on binding of RGD sequences to the platelet receptor alpha IIb beta 3 we have used recombinant DNA techniques to install the RGD sequence into 'presentation scaffolds', small proteins of known 3-D structure chosen to present guest sequences in constrained orientations. Using Escherichia coli expression systems we made sequence variants in which loop residues of the immunoglobulin VL domain REI and of human interleukin-1 beta were replaced (without changing polypeptide length) by the RGD sequence at positions predicted, based on small molecule studies, to orient the RGD moiety into an active conformation. These variants do not compete for fibrinogen binding to alpha IIb beta 3 up to almost 1 mM concentration. Unfolded or proteolytically fragmented forms of these same proteins do compete, however, showing that the RGD sequences in the mutants must be prohibited from binding by constraints imposed by scaffold structure. To suppress the effects of such structural constraints we constructed two sequence variants in which RGD-containing sequences 42-57 or 44-55 from the snake venom platelet antagonist kistrin were inserted (this increasing the length of the loop) into the third complementarity determining loop of REI. Both of these variants compete strongly for fibrinogen binding with IC50s in the nM range. These results, plus data on kistrin-related peptides also presented here, suggest that the molecular scaffold REI is capable of providing to an installed sequence a structural context and conformation beneficial to binding. The results also suggest that in order to bind well to alpha IIb beta 3, RGD sequences in protein ligands must either project significantly from the surface of the scaffold and/or retain a degree of conformational flexibility within the scaffold. Molecular scaffolds like REI should prove useful in the elucidation of structure-function relationships and the discovery of new active sequences, and may also serve as the basis for novel therapeutic agents.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0269-2139
pubmed:author
pubmed:issnType
Print
pubmed:volume
6
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
745-54
pubmed:dateRevised
2005-11-17
pubmed:meshHeading
pubmed-meshheading:8248098-Amino Acid Sequence, pubmed-meshheading:8248098-Base Sequence, pubmed-meshheading:8248098-Binding, Competitive, pubmed-meshheading:8248098-Binding Sites, pubmed-meshheading:8248098-Escherichia coli, pubmed-meshheading:8248098-Fibrinogen, pubmed-meshheading:8248098-Immunoglobulin Light Chains, pubmed-meshheading:8248098-Immunoglobulin Variable Region, pubmed-meshheading:8248098-Interleukin-1, pubmed-meshheading:8248098-Molecular Sequence Data, pubmed-meshheading:8248098-Mutagenesis, pubmed-meshheading:8248098-Oligopeptides, pubmed-meshheading:8248098-Peptides, pubmed-meshheading:8248098-Platelet Membrane Glycoproteins, pubmed-meshheading:8248098-Protein Conformation, pubmed-meshheading:8248098-Protein Folding, pubmed-meshheading:8248098-Recombinant Fusion Proteins, pubmed-meshheading:8248098-Transformation, Bacterial
pubmed:year
1993
pubmed:articleTitle
Strong inhibition of fibrinogen binding to platelet receptor alpha IIb beta 3 by RGD sequences installed into a presentation scaffold.
pubmed:affiliation
Macromolecular Sciences, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406.
pubmed:publicationType
Journal Article