Dataset Information

Mechanistic Basis for the Binding of RGD- and AGDV-Peptides to the Platelet Integrin ?IIb?3.

ABSTRACT: Binding of soluble fibrinogen to the activated conformation of the integrin ?IIb?3 is required for platelet aggregation and is mediated exclusively by the C-terminal AGDV-containing dodecapeptide (?C-12) sequence of the fibrinogen ? chain. However, peptides containing the Arg-Gly-Asp (RGD) sequences located in two places in the fibrinogen A? chain inhibit soluble fibrinogen binding to ?IIb?3 and make substantial contributions to ?IIb?3 binding when fibrinogen is immobilized and when it is converted to fibrin. Here, we employed optical trap-based nanomechanical measurements and computational molecular modeling to determine the kinetics, energetics, and structural details of cyclic RGDFK (cRGDFK) and ?C-12 binding to ?IIb?3. Docking analysis revealed that NMR-determined solution structures of cRGDFK and ?C-12 bind to both the open and closed ?IIb?3 conformers at the interface between the ?IIb ?-propeller domain and the ?3 ?I domain. The nanomechanical measurements revealed that cRGDFK binds to ?IIb?3 at least as tightly as ?C-12. A subsequent analysis of molecular force profiles and the number of peptide-?IIb?3 binding contacts revealed that both peptides form stable bimolecular complexes with ?IIb?3 that dissociate in the 60-120 pN range. The Gibbs free energy profiles of the ?IIb?3-peptide complexes revealed that the overall stability of the ?IIb?3-cRGDFK complex was comparable with that of the ?IIb?3-?C-12 complex. Thus, these results provide a mechanistic explanation for previous observations that RGD- and AGDV-containing peptides are both potent inhibitors of the ?IIb?3-fibrinogen interactions and are consistent with the observation that RGD motifs, in addition to AGDV, support interaction of ?IIb?3 with immobilized fibrinogen and fibrin.

SUBMITTER: Kononova O

PROVIDER: S-EPMC5536102 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Mechanistic Basis for the Binding of RGD- and AGDV-Peptides to the Platelet Integrin αIIbβ3.

Kononova Olga O Litvinov Rustem I RI Blokhin Dmitry S DS Klochkov Vladimir V VV Weisel John W JW Bennett Joel S JS Barsegov Valeri V

Biochemistry 20170322 13

Binding of soluble fibrinogen to the activated conformation of the integrin αIIbβ3 is required for platelet aggregation and is mediated exclusively by the C-terminal AGDV-containing dodecapeptide (γC-12) sequence of the fibrinogen γ chain. However, peptides containing the Arg-Gly-Asp (RGD) sequences located in two places in the fibrinogen Aα chain inhibit soluble fibrinogen binding to αIIbβ3 and make substantial contributions to αIIbβ3 binding when fibrinogen is immobilized and when it is conver ...[more]

PMID: 28277676

Similar Datasets

Project description:The major platelet integrin, glycoprotein IIb-IIIa, binds soluble fibrinogen only after platelet activation. To investigate the mechanism by which platelets convert glycoprotein IIb-IIIa into a functional fibrinogen receptor, we characterized the opening and closing of fibrinogen-binding sites in isolated platelet membranes and compared the regulatory properties of membrane-bound glycoprotein IIb-IIIa with those of the detergent-solubilized receptor. Basal fibrinogen binding to the membranes possessed many of the properties of fibrinogen binding to activated platelets; however, less than 10% of glycoprotein IIb-IIIa in the membranes was capable of binding fibrinogen. Preincubating the membranes with either an activating glycoprotein IIb-IIIa antibody or alpha-chymotrypsin increased fibrinogen binding. In contrast, agents that require intracellular mediators, such as platelet agonists, guanine-nucleotide-binding-protein activators and purified protein kinase C, did not stimulate fibrinogen binding to the membranes, suggesting that cytosolic factor(s) may be required for activation of the receptor in platelets. Occupancy of glycoprotein IIb-IIIa in the membranes with RGD (Arg-Gly-Asp)-containing peptides reversibly exposed neoantigenic epitopes and fibrinogen-binding sites in the receptor. These conformational changes required membrane fixation to be maintained following peptide removal. Similar results were obtained with purified glycoprotein IIb-IIIa incorporated into phospholipid vesicles, indicating that the resting state of the receptor is favoured in these environments. In contrast, when the conformation of detergent-solubilized glycoprotein IIb-IIIa was altered by exposure to RGD-containing peptides, the receptor remained active even after incorporation into phospholipid vesicles. These results demonstrate that platelet membranes are a useful model in which to study the regulation of glycoprotein IIb-IIIa and suggest that the environment surrounding the receptor may have a profound influence on this process.

Project description:Integrin alpha(v)beta(3) plays a significant role in tumor angiogenesis and is a receptor for the extracellular matrix proteins with the exposed arginine-glycine-aspartic (RGD) tripeptide sequence. These include vitronectin, fibronectin, fibrinogen, lamin, collagen, Von Willibrand's factor, osteoponin, and adenovirus particles. Integrin alpha(v)beta(3) is expressed at low levels on epithelial cells and mature endothelial cells, but it is overexpressed on the activated endothelial cells of tumor neovasculature and some tumor cells. The restricted expression of integrin alpha(v)beta(3) during tumor growth, invasion, and metastasis presents an interesting molecular target for both early detection and treatment of rapidly growing solid tumors. Over the past decade, many radiolabeled linear and cyclic RGD peptide antagonists have been evaluated as integrin alpha(v)beta(3)-targeted radiotracers. Significant progress has been made on their use for imaging integrin alpha(v)beta(3)-positive tumors by SPECT or PET. Among the radiotracers evaluated in preclinical tumor-bearing models, [18F]Galacto-RGD (2-[18F]fluoropropanamide c(RGDfK(SAA); SAA = 7-amino-L-glyero-L-galacto-2,6-anhydro-7-deoxyheptanamide) and [18F]-AH111585 are currently under clinical investigation for visualization of integrin alpha(v)beta(3) expression in cancer patients. However, their low tumor uptake, high cost, and lack of preparative modules for routine radiosynthesis will limit their continued clinical application. Thus, there is a continuing need for more efficient integrin alpha(v)beta(3)-targeted radiotracers that are readily prepared from a kit formulation without further postlabeling purification. This article will focus on different approaches to maximize the targeting capability of cyclic RGD peptides and to improve the radiotracer excretion kinetics from noncancerous organs. Improvement of tumor uptake and tumor-to-background ratios is important for early detection of integrin alpha(v)beta(3)-positive tumors and/or noninvasive monitoring of therapeutic efficacy of antiangiogenic therapy.

Dataset Information

Mechanistic Basis for the Binding of RGD- and AGDV-Peptides to the Platelet Integrin ?IIb?3.

Publications

Mechanistic Basis for the Binding of RGD- and AGDV-Peptides to the Platelet Integrin αIIbβ3.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets