Enhanced Local Disorder in a Clinically Elusive von Willebrand Factor Provokes High-Affinity Platelet Clumping

JOURNAL OF MOLECULAR BIOLOGY

卷:429

Article

Tischer, Alexander^1,2  Machha, Venkata R.^1,2  Frontroth, Juan P.³  Brehm, Maria A.⁴  Obser, Tobias⁴  Schneppenheim, Reinhard⁴  Mayne, Leland⁵  Englander, S. Walter⁵  Auton, Matthew^1,2 

[1] Mayo Clin, Dept Internal Med, Div Hematol, Rochester, MN 55905 USA

[2] Mayo Clin, Dept Biochem & Mol Biol, Rochester, MN 55905 USA

[3] Hosp Pediatria, Lab Hemostasis & Trombosis, Serv Hematol & Oncol, Buenos Aires, DF, Argentina

[4] Univ Med Ctr Hamburg Eppendorf, Dept Paediat Haematol & Oncol, Hamburg, Germany

[5] Univ Penn, Perelman Sch Med, Dept Biochem & Biophys, Johnson Res Fdn, Philadelphia, PA 19104 USA

关键词: Von Willebrand factor;    Von Willebrand disease;    hydrogen-deuterium exchange mass spectrometry;    GPlb alpha;   

DOI  :  10.1016/j.jmb.2017.05.013

来源: Elsevier

【 摘 要 】

Mutation of the cysteines forming the disulfide loop of the platelet GPlb alpha adhesive A1 domain of von Willebrand factor (VWF) causes quantitative VWF deficiencies in the blood and von Willebrand disease. We report two cases of transient severe thrombocytopenia induced by DDAVP treatment. Cys1272Trp and Cys1458Tyr mutations identified by genetic sequencing implicate an abnormal gain-of-function phenotype, evidenced by thrombocytopenia, which quickly relapses back to normal platelet counts and deficient plasma VWF. Using surface plasmon resonance, analytical rheology, and hydrogen deuterium exchange mass spectrometry (HXMS), we decipher mechanisms of Al-GPlb alpha-mediated platelet adhesion and resolve dynamic secondary structure elements that regulate the binding pathway. Constrained by the disulfide, conformational selection between weak and tight binding states of A1 takes precedence and drives normal platelet adhesion to VWF. Less restrained through mutation, loss of the disulfide preferentially diverts binding through an induced-fit disease pathway enabling high-affinity GPlba binding and firm platelet adhesion to a partially disordered A1 domain. HXMS reveals a dynamic asymmetry of flexible and ordered regions common to both variants, indicating that the partially disordered A1 lacking the disulfide retains native-like structural dynamics. Both binding mechanisms share common structural and thermodynamic properties, but the enhanced local disorder in the disease state perpetuates high-affinity platelet agglutination, characteristic of type 2B VWD, upon DDAVP-stimulated secretion of VWF leading to transient thrombocytopenia and a subsequent deficiency of plasma VWF, characteristic of type 2A VWD. (C) 2017 Elsevier Ltd. All rights reserved.

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