A Barbourin-albumin Fusion Protein that Is Slowly Cleared In Vivo Retains the Ability to Inhibit Platelet Aggregation In Vitro

Janey A. Marques; Jacob K. George; Ian J. Smith; Varsha Bhakta; William P. Sheffield

doi:10.1055/s-0037-1616148

Thrombosis and Haemostasis, Inhaltsverzeichnis

Thromb Haemost 2001; 86(03): 902-908
DOI: 10.1055/s-0037-1616148

Review Articles

Schattauer GmbH

A Barbourin-albumin Fusion Protein that Is Slowly Cleared In Vivo Retains the Ability to Inhibit Platelet Aggregation In Vitro

Janey A. Marques

¹Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada

,

Jacob K. George

¹Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada

,

Ian J. Smith

¹Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada

,

Varsha Bhakta

¹Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada

,

William P. Sheffield

¹Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada

²Canadian Blood Services, Research and Development, Canada

› Institutsangaben

Abstract

Summary

Barbourin is a 73 amino acid venom protein that inhibits platelet aggregation. Recombinant barbourin (BARH₆), rabbit serum albumin (RSAH₆), and a barbourin-RSA fusion protein (barbourin-linker-albumin; BLAH₆) were secreted from Pichia pastoris yeast, and purified by nickel-chelate affinity chromatography via their C-terminal hexahisti-dine (H₆) tags. BARH₆ and BLAH₆ did not differ in their IC₅₀s for inhibition of platelet aggregation using either human platelets stimulated with thrombin or ADP, or rabbit platelets stimulated with ADP. BARH₆ and BLAH₆ were also effective in inhibiting platelet aggregation in whole blood, and formed complexes with platelet integrin α_IIbβ₃. The terminal catabolic half-life of BLAH₆ approached that of RSAH₆[3.4 ± 0.2 versus 4.0 ± 0.1 days (n = 4 ± SD)], but was substantially increased relative to that of BARH₆ [0.15 ± 0.03 days (n = 3 ± SD)]. Our results suggest that fusion to albumin slows the clearance of barbourin in vivo, while preserving its ability to inhibit platelet aggregation.

Keywords

Barbourin - disintegrins - albumin - clearance

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