HOMOZYGOUS AND HETEROZYGOUS COMPLETE DELETIONS OF THE VON WILLEBRAND FACTOR GENE CODING REGION IN SEVERE VON WILLEBRAND DISEASE AND CARRIERS

1987 ◽  
Author(s):  
K Y Ngo ◽  
D Lynch ◽  
J Gitscher ◽  
N Ciavarella ◽  
Z Ruggeri ◽  
...  
Keyword(s):  
Amino Acid ◽  
Von Willebrand Factor ◽  
Genomic Dna ◽  
Untranslated Region ◽  
Gene Dosage ◽  
Von Willebrand Disease ◽  
Coding Region ◽  
Gene Coding ◽  
Von Willebrand ◽  
Willebrand Factor

Severe von Willebrand disease (vWD) is characterized by undetectable levels of von Willebrand factor (vWF), or trace amounts, in plasma and tissue stores. We have studied the genomic DNA of ten affected individuals from five families with this disorder using two cDNA probes. One probe extended from 175 base pairs of the 5’ untranslated region to the nucleotides encoding amino acid 618 of pro-vWF; the second extended from the nucleotides encoding amino acid 2225 of pro-vWF to 100 bp into the 3’ untranslated region. Three variants of the disorder were identified. Southern blots of restriction endonuclease digests and slot blots of undigested genomic DNA showed complete homozygous deletion of the vWF gene coding region in four affected siblings, three of whom had developed allo-antibodies. Gene dosage analysis performed with slot blots and laser densitometry were consistent with complete heterozygous deletions in both parents. The second variant was characterized by a complete heterozygous deletion of the vWF gene coding region in the propositus and one asymptomatic parent, suggesting that a different type of genetic abnormality was inherited from the other parent and that the patient was doubly heterozygous for distinct genetic abnormalities affecting vWF. In a third variant, no abnormalities could be detected. These techniques should prove useful in identifying carriers of severe vWD and also defining patients at risk of developing allo-antibodies to vWF.

scholarly journals The von willebrand factor domain-mediating botrocetin-induced binding to glycoprotein IB lies between Val449 and Lys728

Blood ◽  
1987 ◽  
Vol 70 (4) ◽  
pp. 985-988 ◽  
Author(s):  
Y Fujimura ◽  
LZ Holland ◽  
ZM Ruggeri ◽  
TS Zimmerman
Keyword(s):  
Monoclonal Antibodies ◽  
Amino Acid ◽  
Von Willebrand Factor ◽  
Amino Acid Residue ◽  
Von Willebrand Disease ◽  
Platelet Glycoprotein ◽  
Tryptic Fragment ◽  
Alternative Agent ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Botrocetin, a component of Bothrops jararaca venom, induces von Willebrand factor (vWF)-dependent platelet agglutination and has been proposed as an alternative agent to ristocetin for evaluating vWF function. However, important differences between the vWF-platelet interactions induced by these two agents have suggested that different regions of vWF and the platelet may be involved in the interactions induced by the two agonists. We have recently demonstrated that binding of vWF to the platelet glycoprotein (GP) Ib receptor, either induced by ristocetin or as occurs spontaneously with asialo-vWF or vWF from IIb von Willebrand disease, is mediated by a domain residing on a 52/48- kilodalton (kD) tryptic fragment of vWF. This fragment extends from amino acid residue Val (449) to Lys (728). We have now found that this 52/48-kD fragment blocks botrocetin-induced binding of vWF to platelets and completely inhibits botrocetin-induced platelet agglutination. These results provide evidence that the vWF domain-mediating, botrocetin-induced platelet agglutination lies within the region delimited by this fragment and is therefore close to or identical with that which mediates ristocetin-induced binding and spontaneous binding of vWF to platelet GPIb. Anti-GPIb monoclonal antibodies also blocked agglutination, which showed that botrocetin, like ristocetin, induces binding of vWF to the GPIb receptor.

scholarly journals The "Normandy" variant of von Willebrand disease: characterization of a point mutation in the von Willebrand factor gene

Blood ◽  
1991 ◽  
Vol 77 (9) ◽  
pp. 1937-1941 ◽  
Author(s):  
C Gaucher ◽  
S Jorieux ◽  
B Mercier ◽  
D Oufkir ◽  
C Mazurier
Keyword(s):  
Amino Acid ◽  
Point Mutation ◽  
Von Willebrand Factor ◽  
Binding Capacity ◽  
Von Willebrand Disease ◽  
Normal Individuals ◽  
New Variant ◽  
Functional Defect ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract We previously reported a functional defect of von Willebrand factor (vWF) in a new variant of von Willebrand disease (vWD) tentatively named vWD “Normandy.” The present work has attempted to characterize the molecular abnormality of this vWF that fails to bind factor VIII (FVIII). The immunopurified vWF from normal and patient's plasma were digested by trypsin and the resulting peptides were compared. The electrophoresis of ““vWF Normandy” showed a shift in the band corresponding to a polypeptide from amino acid 1 to 272. Consequently, we performed the molecular analysis of the portion of the vWF gene of this patient encoding this amino acid sequence. Exons 18–24 were amplified by the use of polymerase chain reaction and their nucleotide sequences corresponding to 1.8 kb were determined. Our analysis showed a point mutation C to T at codon 791, resulting in the substitution of Methionine for Threonine at position 28 of the mature vWF subunit. Because this nucleotide substitution destroyed a Mae II restriction site, this mutation was conveniently sought in various individual DNAs. The patterns obtained were consistent with the homozygous and heterozygous state of this mutation in the patient and in her son, respectively, and with its absence in 28 normal individuals. We conclude that Threonine at position 28 in plasma vWF may be crucial for the conformation and FVIII-binding capacity of its cystine-rich N- terminal domain.

scholarly journals The mutation Arg (53)----Trp causes von Willebrand disease Normandy by abolishing binding to factor VIII. Studies with recombinant von Willebrand factor

Blood ◽  
1992 ◽  
Vol 79 (3) ◽  
pp. 563-567 ◽  
Author(s):  
S Jorieux ◽  
EA Tuley ◽  
C Gaucher ◽  
C Mazurier ◽  
JE Sadler
Keyword(s):  
Amino Acid ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Protein Complex ◽  
Von Willebrand Disease ◽  
Cho Cell ◽  
Fviii Activity ◽  
New Variant ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract von Willebrand factor (vWF) and factor VIII (FVIII) circulate in plasma as a noncovalently linked protein complex. The FVIII/vWF interaction is required for the stabilization of procoagulant FVIII activity. Recently, we reported a new variant of von Willebrand disease (vWD) tentatively named “Normandy,” characterized by plasma vWF that appears to be structurally and functionally normal except that it does not bind FVIII. Three patients from one family were found to be homozygous for a C----T transition at codon 816 converting Arg 53 to Trp in the mature vWF subunit. To firmly establish a causal relationship between this missense mutation and vWD Normandy phenotype, we have characterized the corresponding recombinant mutant vWF(R53W). Expressed in COS-7 cells or CHO cell lines, normal vWF and vWF(R53W) were processed and formed multimers with equal efficiency. However, vWF(R53W) exhibited the same defect in FVIII binding as did plasma vWF from patients with vWD Normandy, confirming that this mutation is responsible for the vWD Normandy phenotype. These results illustrate the importance of Arg 53 of the mature vWF subunit for the binding of FVIII to vWF, and identify an amino acid residue within a disulfide loop not previously known to be involved in this interaction.

scholarly journals The "Normandy" variant of von Willebrand disease: characterization of a point mutation in the von Willebrand factor gene

Blood ◽  
1991 ◽  
Vol 77 (9) ◽  
pp. 1937-1941
Author(s):  
C Gaucher ◽  
S Jorieux ◽  
B Mercier ◽  
D Oufkir ◽  
C Mazurier
Keyword(s):  
Amino Acid ◽  
Point Mutation ◽  
Von Willebrand Factor ◽  
Binding Capacity ◽  
Von Willebrand Disease ◽  
Normal Individuals ◽  
New Variant ◽  
Functional Defect ◽  
Von Willebrand ◽  
Willebrand Factor

We previously reported a functional defect of von Willebrand factor (vWF) in a new variant of von Willebrand disease (vWD) tentatively named vWD “Normandy.” The present work has attempted to characterize the molecular abnormality of this vWF that fails to bind factor VIII (FVIII). The immunopurified vWF from normal and patient's plasma were digested by trypsin and the resulting peptides were compared. The electrophoresis of ““vWF Normandy” showed a shift in the band corresponding to a polypeptide from amino acid 1 to 272. Consequently, we performed the molecular analysis of the portion of the vWF gene of this patient encoding this amino acid sequence. Exons 18–24 were amplified by the use of polymerase chain reaction and their nucleotide sequences corresponding to 1.8 kb were determined. Our analysis showed a point mutation C to T at codon 791, resulting in the substitution of Methionine for Threonine at position 28 of the mature vWF subunit. Because this nucleotide substitution destroyed a Mae II restriction site, this mutation was conveniently sought in various individual DNAs. The patterns obtained were consistent with the homozygous and heterozygous state of this mutation in the patient and in her son, respectively, and with its absence in 28 normal individuals. We conclude that Threonine at position 28 in plasma vWF may be crucial for the conformation and FVIII-binding capacity of its cystine-rich N- terminal domain.

Three Distinct Candidate Point Mutations of the von Willebrand Factor Gene in Four Patients with Type IIA von Willebrand Disease

1992 ◽  
Vol 67 (06) ◽  
pp. 612-617 ◽  
Author(s):  
Isamu Sugiura ◽  
Tadashi Matsushita ◽  
Mitsune Tanimoto ◽  
Isao Takahashi ◽  
Tomio Yamazaki ◽  
...  
Keyword(s):  
Amino Acid ◽  
Von Willebrand Factor ◽  
Cleavage Site ◽  
Point Mutations ◽  
Pcr Amplification ◽  
Von Willebrand Disease ◽  
Missense Mutations ◽  
Allele Specific ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryType IIA von Willebrand disease (vWD) is the most common type II vWD and is characterized by the selective loss of large and intermediate sized multimers. One explanation for this disorder has been postulated to be a qualitative defect in von Willebrand factor (vWF) which results in increased susceptibility to proteolysis at the bond between residues Tyr842 and Met843. Four missense mutations that may cause type IIA vWD have recently been identified near the cleavage site. We analyzed the molecular basis for type IIA vWD in six patients. A 512 bp DNA sequence spanning the proteolytic cleavage site was targeted for PCR amplification and sequencing. We exploited a difference in restriction sites between the vWF gene and the pseudogene and have designed allele-specific oligomer used with PCR to distinguish these two genes. Three candidate missense mutations; Ser743 (TCG) → Leu (TTG), Leu799 (CTG) → Pro (CCG), and Arg834 (CGG) → Trp (TGG) were identified in 4 out of 6 patients. The amino acid substitution at Arg834 has been reported previously, but the other substitutions at Ser743 and Leu799 are novel candidate mutations locating 99 and 43 amino acids to the N-terminal side of the cleavage site, respectively. Our results indicate that amino acid substitutions located relatively distant from the cleavage site may also be involved in type IIA vWD.

scholarly journals Duplication of a methionine within the glycoprotein Ib binding domain of von Willebrand factor detected by denaturing gradient gel electrophoresis in a patient with type IIB von Willebrand disease

Blood ◽  
1991 ◽  
Vol 78 (7) ◽  
pp. 1738-1743 ◽  
Author(s):  
AS Ribba ◽  
JM Lavergne ◽  
BR Bahnak ◽  
A Derlon ◽  
G Pietu ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Binding Domain ◽  
Von Willebrand Disease ◽  
Functional Domains ◽  
Gene Coding ◽  
Gradient Gel Electrophoresis ◽  
Von Willebrand ◽  
Willebrand Factor

von Willebrand disease (vWD) type IIB is characterized by an increased reactivity of von Willebrand factor (vWF) with platelets and a lack of large multimers. Exon 28 of the vWF gene encodes for functional domains involved in the binding of vWF to GPIb, and it is presumed that the defects in type IIB vWD lie within or adjacent to these functional domains. We screened overlapping DNA fragments generated by the polymerase chain reaction (PCR) that spanned the 1,379 bp of exon 28 of a type IIB vWD patient using denaturing gradient gel electrophoresis (DGGE). To increase the power of DGGE to detect base changes, we used the PCR to attach a G + C-rich sequence. In the type IIB patient, a DNA fragment at the 5′ end of exon 28 demonstrated homoduplex and heteroduplex complexes after DGGE, a pattern characteristic of heterozygous genes after melting and reannealing during the PCR. Sequencing of the cloned insert from the patient showed a duplication of an ATG in one gene coding for a Met at amino acids 540 to 541 in the mature vWF subunit. This duplication leads to three consecutive methionines in the patient's sequence. The duplicated Met resides within a disulfide bond loop proposed to be important in the function of the GPIb binding domain of vWF. The patient's nephew, who also has type IIB vWD, showed the same duplicated codon, linking the defect to the abnormal phenotype in this family. These nucleotide changes were not found in 100 chromosomes analyzed either by DGGE or hybridization with an allele specific oligonucleotide containing the duplicated ATG codon. In addition, the same oligonucleotide hybridized only to DNA from type IIB vWD individuals and not to DNA from normal members of the family. Therefore, we conclude that this duplicated Met modifies the GPIb binding domain of vWF and causes type IIB vWD in this family.

scholarly journals Type 2M von Willebrand Disease: F606I and I662F Mutations in the Glycoprotein Ib Binding Domain Selectively Impair Ristocetin- but not Botrocetin-Mediated Binding of von Willebrand Factor to Platelets

Blood ◽  
1998 ◽  
Vol 91 (5) ◽  
pp. 1572-1581 ◽  
Author(s):  
Cheryl A. Hillery ◽  
David J. Mancuso ◽  
J. Evan Sadler ◽  
Jay W. Ponder ◽  
Mary A. Jozwiak ◽  
...  
Keyword(s):  
Amino Acid ◽  
Missense Mutation ◽  
Von Willebrand Factor ◽  
Amino Acid Substitution ◽  
Von Willebrand Disease ◽  
Glycoprotein Ib ◽  
Collagen Binding ◽  
Platelet Binding ◽  
Von Willebrand ◽  
Willebrand Factor

Abstractvon Willebrand disease (vWD) is a common, autosomally inherited, bleeding disorder caused by quantitative and/or qualitative deficiency of von Willebrand factor (vWF). We describe two families with a variant form of vWD where affected members of both families have borderline or low vWF antigen levels, normal vWF multimer patterns, disproportionately low ristocetin cofactor activity, and significant bleeding symptoms. Whereas ristocetin-induced binding of plasma vWF from affected members of both families to fixed platelets was reduced, botrocetin-induced platelet binding was normal. The sequencing of genomic DNA identified unique missense mutations in each family in the vWF exon 28. In Family A, a missense mutation at nucleotide 4105T → A resulted in a Phe606Ile amino acid substitution (F606I) and in Family B, a missense mutation at nucleotide 4273A → T resulted in an Ile662Phe amino acid substitution (I662F). Both mutations are within the large disulfide loop between Cys509 and Cys695 in the A1 domain that mediates vWF interaction with platelet glycoprotein Ib. Expression of recombinant vWF containing either F606I or I662F mutations resulted in mutant recombinant vWF with decreased ristocetin-induced platelet binding, but normal multimer structure, botrocetin-induced platelet binding, collagen binding, and binding to the conformation-sensitive monoclonal antibody, AvW-3. Both mutations are phenotypically distinct from the previously reported variant type 2MMilwaukee-1 because of the presence of normal botrocetin-induced platelet binding, collagen binding, and AvW-3 binding, as well as the greater frequency and intensity of clinical bleeding. When the reported type 2M mutations are mapped on the predicted three-dimensional structure of the A1 loop of vWF, the mutations cluster in one region that is distinct from the region in which the type 2B mutations cluster.

scholarly journals Type 2M von Willebrand Disease: F606I and I662F Mutations in the Glycoprotein Ib Binding Domain Selectively Impair Ristocetin- but not Botrocetin-Mediated Binding of von Willebrand Factor to Platelets

Blood ◽  
1998 ◽  
Vol 91 (5) ◽  
pp. 1572-1581 ◽  
Author(s):  
Cheryl A. Hillery ◽  
David J. Mancuso ◽  
J. Evan Sadler ◽  
Jay W. Ponder ◽  
Mary A. Jozwiak ◽  
...  
Keyword(s):  
Amino Acid ◽  
Missense Mutation ◽  
Von Willebrand Factor ◽  
Amino Acid Substitution ◽  
Von Willebrand Disease ◽  
Glycoprotein Ib ◽  
Collagen Binding ◽  
Platelet Binding ◽  
Von Willebrand ◽  
Willebrand Factor

von Willebrand disease (vWD) is a common, autosomally inherited, bleeding disorder caused by quantitative and/or qualitative deficiency of von Willebrand factor (vWF). We describe two families with a variant form of vWD where affected members of both families have borderline or low vWF antigen levels, normal vWF multimer patterns, disproportionately low ristocetin cofactor activity, and significant bleeding symptoms. Whereas ristocetin-induced binding of plasma vWF from affected members of both families to fixed platelets was reduced, botrocetin-induced platelet binding was normal. The sequencing of genomic DNA identified unique missense mutations in each family in the vWF exon 28. In Family A, a missense mutation at nucleotide 4105T → A resulted in a Phe606Ile amino acid substitution (F606I) and in Family B, a missense mutation at nucleotide 4273A → T resulted in an Ile662Phe amino acid substitution (I662F). Both mutations are within the large disulfide loop between Cys509 and Cys695 in the A1 domain that mediates vWF interaction with platelet glycoprotein Ib. Expression of recombinant vWF containing either F606I or I662F mutations resulted in mutant recombinant vWF with decreased ristocetin-induced platelet binding, but normal multimer structure, botrocetin-induced platelet binding, collagen binding, and binding to the conformation-sensitive monoclonal antibody, AvW-3. Both mutations are phenotypically distinct from the previously reported variant type 2MMilwaukee-1 because of the presence of normal botrocetin-induced platelet binding, collagen binding, and AvW-3 binding, as well as the greater frequency and intensity of clinical bleeding. When the reported type 2M mutations are mapped on the predicted three-dimensional structure of the A1 loop of vWF, the mutations cluster in one region that is distinct from the region in which the type 2B mutations cluster.

An amino acid polymorphism in von Willebrand factor correlates with increased susceptibility to proteolysis by ADAMTS13

Blood ◽  
2004 ◽  
Vol 103 (3) ◽  
pp. 941-947 ◽  
Author(s):  
Derrick John Bowen ◽  
Peter William Collins
Keyword(s):  
Amino Acid ◽  
Von Willebrand Factor ◽  
Von Willebrand Disease ◽  
Type I ◽  
Adamts13 Activity ◽  
Amino Acid Polymorphism ◽  
A Prospective Study ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Increased Susceptibility

Abstract The hypothesis that increased ADAMTS13 (a disintegrin and metalloprotease with thrombospondin repeats) activity or increased susceptibility of von Willebrand factor (VWF) to proteolysis by ADAMTS13 may underlie type I von Willebrand disease (VWD) in some patients was investigated. Plasma from 4 patients with type I VWD was cryoprecipitated. ADAMTS13 activity in the VWF-poor cryodepleted fraction was assessed by incubation with purified VWF; susceptibility to proteolysis of the VWF in the VWF-rich cryoprecipitate was assessed by incubation with a normal, group O cryodepleted plasma. ADAMTS13 activity was similar in all 4 type I VWD cryodepleted plasmas and comparable to a normal control plasma. In contrast, the VWF of one patient showed increased susceptibility to proteolysis by ADAMTS13. Investigation of additional family members indicated that increased susceptibility was heritable, but it did not track uniquely with type I VWD. Sequence analysis of VWF exon 28 indicated that increased susceptibility to proteolysis tracked with the “G” allele of the A/G polymorphism at position 24/1282, encoding the amino acid polymorphism Tyr/Cys1584 (“G” = Cys1584). A prospective study of 200 individuals yielded 2 Tyr/Cys1584 heterozygotes; for both, plasma VWF showed increased susceptibility to proteolysis. The finding that an amino acid polymorphism in VWF may influence susceptibility to ADAMTS13 has potentially significant implications in diverse areas. (Blood. 2004;103:941-947)

A Standard Nomenclature for von Willebrand Factor Gene Mutations and Polymorphisms

2001 ◽  
Vol 85 (05) ◽  
pp. 929-931 ◽  
Author(s):  
J. C. J. Eikenboom ◽  
D. Ginsburg ◽  
L. Hilbert ◽  
C. Mazurier ◽  
I. R. Peake ◽  
...  
Keyword(s):  
Amino Acid ◽  
Von Willebrand Factor ◽  
Gene Mutations ◽  
Von Willebrand Disease ◽  
Amino Acid Code ◽  
Cdna Nucleotide Sequence ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Cdna Nucleotide ◽  
Single Sequence

SummaryExamination of the entire von Willebrand factor (VWF) gene for mutations, particularly in types 1 and 3 von Willebrand disease (VWD) is becoming more widely practised. The sequence of the entire VWF gene will soon be compiled as a single sequence. For these reasons, a clearly defined nomenclature to use for numbering the VWF nucleotide and amino acid sequence is required.The following recommendations are made for VWF numbering. VWF cDNA nucleotide sequence should be numbered from the A of the initiator ATG as the +1 position. Genomic DNA should be prefixed with a “g” and also numbered from this position. Amino acid (aa) numbering should be from the initiator methionine as the +1 position with sequential numbering of aa throughout VWF. To avoid confusion with previously used numbering schemes for mature VWF, which started from serine 764 of pre-pro VWF, the use of the single letter amino acid code is recommended.

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