Five Novel Mutations of the Protein S Active Gene (PROS 1) in 8 Norman Families

1996 ◽  
Vol 75 (03) ◽  
pp. 437-444 ◽  
Author(s):  
Jérôme Duchemin ◽  
Jeanne-Yvonne Borg ◽  
Delphine Borgel ◽  
Marc Vasse ◽  
Hervé Lévèque ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Molecular Basis ◽  
Protein S ◽  
Type I ◽  
Factor V ◽  
Novel Mutations ◽  
Nucleotide Deletion ◽  
Active Gene ◽  
Sequence Variations ◽  
Gradient Gel Electrophoresis

SummaryTo further elucidate the molecular basis for hereditary thrombophilia, we screened the protein S active gene in 11 families with type I deficiency, using a strategy based on denaturating gradient gel electrophoresis (DGGE) of all the coding sequences. Fragments with an abnormal DGGE pattern were sequenced, and 5 novel mutations were identified in 8 families. The mutations were a 7-nucleotide deletion in exon II, a 4-nucleotide deletion in exon III, a T insertion in exon VII, a C to T transition transforming Leu 259 into Pro and a T to C transition transforming Cys 625 into Arg in 4 families. These mutations were the only sequence variations found in the propositus’ gene exons and co-segregated with the plasma phenotype. A total of 28 members of these 8 families were heterozygous for one of the 5 mutations. Twenty-four (58,5%) of the 41 deficient subjects over 18 years of age had clinical thrombophilia, whereas the 13 subjects under 18 were asymptomatic. Of the 28 subjects, 6 (21,5%) were also found to bear the factor V Arg 506 Gin mutation.

Five novel mutations in the gene for human blood coagulation factor V associated with type I factor V deficiency

Blood ◽  
2001 ◽  
Vol 98 (2) ◽  
pp. 358-367 ◽  
Author(s):  
Richard van Wijk ◽  
Karel Nieuwenhuis ◽  
Marijke van den Berg ◽  
Eric G. Huizinga ◽  
Brenda B. van der Meijden ◽  
...  
Keyword(s):  
Missense Mutation ◽  
Base Pair ◽  
Molecular Basis ◽  
Coagulation Factor ◽  
Coagulation Disorder ◽  
Type I ◽  
Factor V ◽  
Novel Mutations ◽  
Base Pair Deletion ◽  
Coagulation Factor V

Coagulation factor V (FV) plays an important role in maintaining the hemostatic balance in both the formation of thrombin in the procoagulant pathway as well as in the protein C anticoagulant pathway. FV deficiency is a rare bleeding disorder with variable phenotypic expression. Little is known about the molecular basis underlying this disease. This study identified 5 novel mutations associated with FV deficiency in 3 patients with severe FV deficiency but different clinical expression and 2 unaffected carriers. Four mutations led to a premature termination codon either by a nonsense mutation (single-letter amino acid codes): A1102T, K310Term. (FV Amersfoort) and C2491T, Q773Term. (FV Casablanca) or a frameshift: an 8–base pair deletion between nucleotides 1130 and 1139 (FV Seoul1) and a 1–base pair deletion between nucleotides 4291 and 4294 (FV Utrecht). One mutation was a novel missense mutation: T1927C, C585R (FV Nijkerk), resulting in the absence of mutant protein despite normal transcription to RNA. Most likely, an arginine at this position disrupts the hydrophobic interior of the FV A2 domain. The sixth detected mutation was a previously reported missense mutation: A5279G, Y1702C (FV Seoul2). In all cases, the presence of the mutation was associated with type I FV deficiency. Identifying the molecular basis of mutations underlying this rare coagulation disorder will help to obtain more insight into the mechanisms involved in the variable clinical phenotype of patients with FV deficiency.

scholarly journals Molecular basis of protein S deficiency in three families also showing independent inheritance of factor V leiden

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1700-1707 ◽  
Author(s):  
NJ Beauchamp ◽  
ME Daly ◽  
PC Cooper ◽  
M Makris ◽  
FE Preston ◽  
...  
Keyword(s):  
Molecular Basis ◽  
Factor V Leiden ◽  
Protein S ◽  
Additional Risk Factor ◽  
Type I ◽  
Factor V ◽  
Protein S Deficiency ◽  
Free Protein ◽  
S Deficiency ◽  
Fv Leiden

The molecular basis of type I or III Protein S deficiency has been investigated in three kindred also showing independent inheritance of factor V (FV) Leiden. A T to C transition in codon 570 (Met-->Thr) was identified in the propositi and shown to segregate with protein S deficiency in all but one of the affected members of two kindred. This individual was heterozygous for a second transition (C to T) causing substitution of serine 624 by leucine. A second member of the same family, with markedly reduced free protein S levels when compared with affected relatives, was heterozygous for both mutations. Haplotype analysis of individuals with the mutated ATG570ACG allele in the two kindred suggested they may have been related by a common ancestor. A G to A transition resulting in substitution of cysteine 145 by tyrosine was detected in the third kindred. All mutations are believed to interfere with protein S binding to C4b-binding protein resulting in reduced free protein S levels. Of the five individuals studied who had experienced thrombotic events, three had combined protein S deficiency and FV Leiden reemphasising the importance of FV Leiden as an additional risk factor for thrombosis in protein S deficiency.

scholarly journals Molecular basis of protein S deficiency in three families also showing independent inheritance of factor V leiden

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1700-1707 ◽  
Author(s):  
NJ Beauchamp ◽  
ME Daly ◽  
PC Cooper ◽  
M Makris ◽  
FE Preston ◽  
...  
Keyword(s):  
Molecular Basis ◽  
Factor V Leiden ◽  
Protein S ◽  
Additional Risk Factor ◽  
Type I ◽  
Factor V ◽  
Protein S Deficiency ◽  
Free Protein ◽  
S Deficiency ◽  
Fv Leiden

Abstract The molecular basis of type I or III Protein S deficiency has been investigated in three kindred also showing independent inheritance of factor V (FV) Leiden. A T to C transition in codon 570 (Met-->Thr) was identified in the propositi and shown to segregate with protein S deficiency in all but one of the affected members of two kindred. This individual was heterozygous for a second transition (C to T) causing substitution of serine 624 by leucine. A second member of the same family, with markedly reduced free protein S levels when compared with affected relatives, was heterozygous for both mutations. Haplotype analysis of individuals with the mutated ATG570ACG allele in the two kindred suggested they may have been related by a common ancestor. A G to A transition resulting in substitution of cysteine 145 by tyrosine was detected in the third kindred. All mutations are believed to interfere with protein S binding to C4b-binding protein resulting in reduced free protein S levels. Of the five individuals studied who had experienced thrombotic events, three had combined protein S deficiency and FV Leiden reemphasising the importance of FV Leiden as an additional risk factor for thrombosis in protein S deficiency.

THROMBOPHILIA:ITS CAUSES AND A ROUGH ESTIMATE OF ITS PREVALENCE

1987 ◽  
Author(s):  
E Briët ◽  
L Engesser ◽  
E J P Brommer ◽  
A W Broekmans ◽  
R M Bertina
Keyword(s):  
Plasminogen Activator ◽  
Protein C ◽  
Antithrombin Iii ◽  
Plasminogen Activator Inhibitor ◽  
Protein S ◽  
Factor Vii ◽  
Type I ◽  
Factor V ◽  
Protein C Deficiency ◽  
Familial Cases

Idiopathic venous thrombosis and embolism have gained widespread interest since the discovery that, deficiencies of antithrombin III, protein C, and protein S are associated with familial venous thrombophilia. The purpose of our study was to obtain an estimate of the prevalence of this syndrome and to establish the etiology in as many cases as possible.We collaborated with specialists from 37 Dutch hospitals, covering about 10% of the Dutch population. A history as well as blood samples were obtained from 113 unrelated cases with familial thrombophilia and from 90 isolated cases. Assuming that each proband in a family with thrombophilia has an average of four affected relatives, a rough estimate of the prevalence of familial thrombophilia in The Netherlands is 40 cases per 100.000. The prevalence of non-familial thrombophilia is probably lower.In 35 out of the 113 familial cases we established a diagnosis of hereditary antithrombin III deficiency (n=5), protein C deficiency (type I: n=9; type II: n=4), protein S deficiency (n=15) and dysfibrinogenemia (n=2). In 36 cases we found no abnormality at all and in the remaining 42 cases abnormalities were found in one or more of the following: heparin cofactor II, factor V, factor VII, factor VIII, von Willebrand factor, plasminogen, tissue plasminogen activator, plasminogen activator inhibitor, alpha 2 antiplasmin and histidine rich glycoprotein. In most of these cases, however, the hereditary nature of the abnormalities could not be demonstrated and the causal relationships remain to be established.In the 90 isolated cases, we diagnosed hereditary deficiencies of anti thrombin III, protein C and protein S each in one case and a lupus anticoagulant in two cases. In 54 cases no abnormality was found and in the remaining 31 cases various abnormalities were found in one or more of the proteins mentioned above.We conclude that the syndrome of thrombophilia is not rare but its true prevalence needs to be established by more rigorous means. An etiological diagnosis can be made with confidence in only one third of the familial cases and in less than 10 percent of the isolated cases.

scholarly journals Identification of 15 different candidate causal point mutations and three polymorphisms in 19 patients with protein S deficiency using a scanning method for the analysis of the protein S active gene

Blood ◽  
1995 ◽  
Vol 85 (1) ◽  
pp. 130-138 ◽  
Author(s):  
S Gandrille ◽  
D Borgel ◽  
V Eschwege-Gufflet ◽  
M Aillaud ◽  
M Dreyfus ◽  
...  
Keyword(s):  
Denaturing Gradient Gel Electrophoresis ◽  
Point Mutations ◽  
Protein S ◽  
Melting Behavior ◽  
Protein S Deficiency ◽  
Scanning Method ◽  
Active Gene ◽  
S Deficiency ◽  
Sequence Variations ◽  
Deficiency Type

To screen for point mutations causing protein S deficiency, we used a sequence of techniques specifically for the study of the protein S active gene, PS alpha. This strategy comprises amplification of exons and intron/exon junctions by means of the polymerase chain reaction (PCR) and electrophoresis of the amplified fragments in polyacrylamide gel containing a gradient of denaturing agents (denaturing gradient gel electrophoresis). Only fragments with altered melting behavior are sequenced after asymmetric PCR. Beside the frequent polymorphism already described on Pro 626, we detected 18 different sequence variations by studying exons II, IV, V, VIII, X, and XV in 19 of 100 consecutive patients with protein S deficiency. Fifteen were candidate causal mutations, 4 of which were associated with a qualitative deficiency (type IIa or IIb). The remaining three sequence variations were probably polymorphisms.

Three novel mutations of the NF1 gene detected by temperature gradient gel electrophoresis of exons 5 and 8

1996 ◽  
Vol 17 (10) ◽  
pp. 1559-1563 ◽  
Author(s):  
Denise Horn ◽  
Peter N. Robinson ◽  
Annett Böddrich ◽  
Annegret Buske ◽  
Sigrid Tinschert ◽  
...  
Keyword(s):  
Temperature Gradient ◽  
Gel Electrophoresis ◽  
Novel Mutations ◽  
Nf1 Gene ◽  
Gradient Gel Electrophoresis ◽  
Temperature Gradient Gel Electrophoresis

scholarly journals Detection of Mitochondrial DNA Mutations by Temporal Temperature Gradient Gel Electrophoresis

1999 ◽  
Vol 45 (8) ◽  
pp. 1162-1167 ◽  
Author(s):  
Tian-Jian Chen ◽  
Richard G Boles ◽  
Lee-Jun C Wong
Keyword(s):  
Mitochondrial Dna ◽  
Temperature Gradient ◽  
Gel Electrophoresis ◽  
Direct Sequencing ◽  
Polymorphism Analysis ◽  
Mtdna Mutations ◽  
Sequence Variations ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
Temperature Gradient Gel Electrophoresis

Abstract Background: A unique requirement for the molecular diagnosis of mitochondrial DNA (mtDNA) disorders is the ability to detect heteroplasmic mtDNA mutations and to distinguish them from homoplasmic sequence variations before further testing (e.g., sequencing) is performed. We evaluated the potential utility of temporal temperature gradient gel electrophoresis (TTGE) for these purposes in patients with suspected mtDNA mutations. Methods: DNA samples were selected from patients with known mtDNA mutations and patients suspected of mtDNA disorders without detectable mutations by routine analysis. Six regions of mtDNA were PCR amplified and analyzed by TTGE. Electrophoresis was carried out at 145 V with a constant temperature increment of 1.2 °C/h. Mutations were identified by direct sequencing of the PCR products and confirmed by PCR/allele-specific oligonucleotide or PCR/restriction fragment length polymorphism analysis. Results: In the experiments using patient samples containing various amounts of mutant mtDNA, TTGE detected as little as 4% mutant heteroplasmy and identified heteroplasmy in the presence of a homoplasmic polymorphism. In 109 specimens with 15 different known mutations, TTGE detected the presence of all mutations and distinguished heteroplasmic mutations from homoplasmic polymorphisms. When 11% of the mtDNA genome was analyzed by TTGE in 104 patients with clinically suspected mitochondrial disorders, 7 cases of heteroplasmy (≈7%) were detected. Conclusions: TTGE distinguishes heteroplasmic mutation from homoplasmic polymorphisms and appears to be a sensitive tool for detection of sequence variations and heteroplasmy in patients suspected of having mtDNA disorders.

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