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Titolo:
STRONG INHIBITION OF FIBRINOGEN BINDING TO PLATELET RECEPTOR -ALPHA-IIB-BETA-3 BY RGD SEQUENCES INSTALLED INTO A PRESENTATION SCAFFOLD
Autore:
LEE G; CHAN W; HURLE MR; DESJARLAIS RL; WATSON F; SATHE GM; WETZEL R;
Indirizzi:
SMITHKLINE BEECHAM PHARMACEUT,DEPT MACROMOLEC SCI,709 SWEDELAND RD KING OF PRUSSIA PA 19406 SMITHKLINE BEECHAM PHARMACEUT,DEPT MACROMOLEC SCI,709 SWEDELAND RD KING OF PRUSSIA PA 19406 SMITHKLINE BEECHAM PHARMACEUT,DEPT MOLEC GENET KING OF PRUSSIA PA 19406 SMITHKLINE BEECHAM PHARMACEUT,DEPT PHYS & STRUCT CHEM KING OF PRUSSIAPA 19406
Titolo Testata:
Protein engineering
fascicolo: 7, volume: 6, anno: 1993,
pagine: 745 - 754
SICI:
0269-2139(1993)6:7<745:SIOFBT>2.0.ZU;2-8
Fonte:
ISI
Lingua:
ENG
Soggetto:
GLYCOPROTEIN-IIB-IIIA; JONES PROTEIN REI; ARG-GLY-ASP; ESCHERICHIA-COLI; AGGREGATION INHIBITOR; CELL-ADHESION; 3-DIMENSIONAL STRUCTURE; DISULFIDE BOND; IMMUNOGLOBULIN LIGHT; SECONDARY STRUCTURE;
Keywords:
FIBRONECTIN; IMMUNOGLOBULIN LIGHT CHAIN VARIABLE DOMAIN; INTERLEUKIN-1-BETA; KISTRIN;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
68
Recensione:
Indirizzi per estratti:
Citazione:
G. Lee et al., "STRONG INHIBITION OF FIBRINOGEN BINDING TO PLATELET RECEPTOR -ALPHA-IIB-BETA-3 BY RGD SEQUENCES INSTALLED INTO A PRESENTATION SCAFFOLD", Protein engineering, 6(7), 1993, pp. 745-754

Abstract

In order to probe the structural constraints on binding of RGD sequences to the platelet receptor alpha(IIb)beta3 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 V(L) domain REI and of human interleukin-1beta 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)beta3 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 - 55from 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)beta3, RGD sequences in protein ligands must either project significantly from the surface of the scaffold and/or retain a degree of conformational flexibilitywithin the scaffold. Molecular scaffolds like REI should prove usefulin the elucidation of structure-function relationships and the discovery of new active sequences, and may also serve as the basis for noveltherapeutic agents.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 21/09/20 alle ore 19:03:56