scholarly journals Antithrombin-Gly 424 Arg: a novel point mutation responsible for type 1 antithrombin deficiency and neonatal thrombosis

Blood ◽  
1994 ◽  
Vol 83 (1) ◽  
pp. 146-151 ◽  
Author(s):  
K Jochmans ◽  
W Lissens ◽  
R Vervoort ◽  
S Peeters ◽  
M De Waele ◽  
...  
Keyword(s):  
Point Mutation ◽  
Direct Sequencing ◽  
Mutant Gene ◽  
Single Strand Conformation Polymorphism ◽  
Molecular Defect ◽  
Polymorphism Analysis ◽  
Functional Protein ◽  
Antithrombin Deficiency ◽  
Increased Risk

Abstract Inherited type 1 antithrombin (AT) III deficiency is characterized by a decrease of immunoreactive and functional protein levels to about 50%. The disorder is associated with a significantly increased risk of thromboembolism. We have investigated the molecular basis of type 1 AT deficiency in a Belgian family. The diagnosis of the disease was primarily made in a newborn girl with unusually severe thrombotic complications. Using the polymerase chain reaction and single-strand conformation polymorphism analysis, followed by direct sequencing of AT gene fragments, we identified a novel point mutation in exon 6. We detected a G to C substitution in the first position of codon 424 leading to a glycine to arginine substitution. The modification at this highly conserved position in the serine protease inhibitor gene family probably leads to an unstable mutant-gene product. The mutation creates a unique restriction site for the enzyme Hha I in exon 6. This change permitted a rapid and accurate screening of the kindred with identification of the molecular defect in five other family members.

scholarly journals Antithrombin-Gly 424 Arg: a novel point mutation responsible for type 1 antithrombin deficiency and neonatal thrombosis

Blood ◽  
1994 ◽  
Vol 83 (1) ◽  
pp. 146-151 ◽  
Author(s):  
K Jochmans ◽  
W Lissens ◽  
R Vervoort ◽  
S Peeters ◽  
M De Waele ◽  
...  
Keyword(s):  
Point Mutation ◽  
Direct Sequencing ◽  
Mutant Gene ◽  
Single Strand Conformation Polymorphism ◽  
Molecular Defect ◽  
Polymorphism Analysis ◽  
Functional Protein ◽  
Antithrombin Deficiency ◽  
Increased Risk

Inherited type 1 antithrombin (AT) III deficiency is characterized by a decrease of immunoreactive and functional protein levels to about 50%. The disorder is associated with a significantly increased risk of thromboembolism. We have investigated the molecular basis of type 1 AT deficiency in a Belgian family. The diagnosis of the disease was primarily made in a newborn girl with unusually severe thrombotic complications. Using the polymerase chain reaction and single-strand conformation polymorphism analysis, followed by direct sequencing of AT gene fragments, we identified a novel point mutation in exon 6. We detected a G to C substitution in the first position of codon 424 leading to a glycine to arginine substitution. The modification at this highly conserved position in the serine protease inhibitor gene family probably leads to an unstable mutant-gene product. The mutation creates a unique restriction site for the enzyme Hha I in exon 6. This change permitted a rapid and accurate screening of the kindred with identification of the molecular defect in five other family members.

The Molecular Basis of Antithrombin Deficiency in Belgian and Dutch Families

1998 ◽  
Vol 80 (09) ◽  
pp. 376-381 ◽  
Author(s):  
W. Lissens ◽  
S. Seneca ◽  
P. Capel ◽  
B. Chatelain ◽  
P. Meeus ◽  
...  
Keyword(s):  
Molecular Basis ◽  
Gene Deletion ◽  
Direct Sequencing ◽  
Genetic Defect ◽  
Polymorphism Analysis ◽  
Type I ◽  
Type Ii ◽  
Antithrombin Deficiency ◽  
Venous Thromboembolic Events ◽  
At Deficiency

SummaryThe molecular basis of hereditary antithrombin (AT) deficiency has been investigated in ten Belgian and three Dutch unrelated kindreds. Eleven of these families had a quantitative or type I AT deficiency, with a history of major venous thromboembolic events in different affected members. In the other two families a qualitative or type II AT deficiency was occasionally diagnosed.DNA studies of the AT gene were performed, using polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis, followed by direct sequencing of the seven exons and intronexon junction regions. Six novel point mutations were identified: four missense, one nonsense mutation and a single nucleotide deletion near the reactive site, causing a frameshift with premature translation termination. In two kindreds the underlying genetic defect was caused by a whole gene deletion, known as a rare cause of AT deficiency. In these cases, Southern blot and polymorphism analysis of different parts of the AT gene proved useful for diagnosis. In another kindred a partial gene deletion spanning 698 basepairs could precisely be determined to a part of intron 3B and exon 4. In two type I and in both type II AT deficient families a previously reported mutation was identified. In all cases, the affected individuals were heterozygous for the genetic defect.

scholarly journals Molecular defect in coagulation factor XFriuli results from a substitution of serine for proline at position 343

Blood ◽  
1991 ◽  
Vol 77 (2) ◽  
pp. 317-323 ◽  
Author(s):  
HL James ◽  
A Girolami ◽  
DS Fair
Keyword(s):  
Point Mutation ◽  
Serine Proteases ◽  
Direct Sequencing ◽  
Coagulation Factor ◽  
Amino Acid Position ◽  
Molecular Defect ◽  
Nucleotide Position ◽  
Factor X ◽  
Gene Coding ◽  
X Gene

Abstract Our previous findings suggested that coagulation factor XFriuli could be functionally defective owing to a point mutation in the portion of the factor X gene coding for the fully activated heavy chain. To verify the existence of this postulated change, we analyzed all eight exons of the normal and Friuli factor X gene. Each exon was amplified from genomic DNA using the polymerase chain reaction and cloned into the plasmid pUC19. The amplified DNA inserts were subjected to direct sequencing by the dideoxy chain termination method with forward and reverse oligonucleotide sequencing primers. A point mutation (C to T transition at nucleotide position 19,297) that results in coding for serine (TCC) in place of proline (CCC) at amino acid position 343 was found. This substitution involves a highly conserved proline residue oriented spatially close to both the cleavage site of the zymogen and the active site of the enzyme and explains the previous observations of discrete biochemical and functional differences between factor XFriuli and normal factor X. The mutation abolished an HgiCI restriction site present in the normal factor X gene, and this change constitutes the basis for a convenient method for screening individuals carrying this molecular defect. Proline343 is in conserved region 5 of the serine protease superfamily to which factor X belongs and is part of a 14- residue L*****P******C motif that occurs in at least 16 other enzymes. Computer analysis suggests that the motif may be an essential aspect of conformational features important to functional properties of factor X as well as other serine proteases.

GATA1 Mutations in Newborns with Down Syndrome.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 641-641
Author(s):  
Claudio Sandoval ◽  
Sharon R. Pine ◽  
Charlotte Druschel ◽  
Somasundaram Jayabose ◽  
Qianxu Guo ◽  
...  
Keyword(s):  
New York ◽  
Down Syndrome ◽  
Single Strand Conformation Polymorphism ◽  
Protein Product ◽  
Polymorphism Analysis ◽  
Exon 2 ◽  
Blood Spots ◽  
Increased Risk ◽  
Significant Difference ◽  
Gata1 Mutation

Abstract Somatic mutations of the GATA1 gene have been detected in almost all cases of Down syndrome (DS)-associated acute megakaryoblastic leukemia (AMKL) and transient leukemia (TL). There is emerging evidence that the protein product of GATA1 mutations, GATA1s, directly contributes to leukemogenesis. Although an in utero origin of GATA1 mutations is established, a comprehensive study using a large number of cases is required to determine the overall incidence and clinical relevance of GATA1 mutations in DS newborns. We screened 575 DS infants born between January 1997 and December 1999 for GATA1 exon 2 mutations by single-strand conformation polymorphism analysis of PCR products. We used Gunthrie cards obtained from the New York Congenital Malformation Registry. Registry data was blinded until after the GATA1 mutation analyses were completed. Twenty-eight (4.9%) infants were identified as having a GATA1 mutation. There was no significant difference in the frequency of GATA1 mutations based on gender or maternal average age (p = 0.93 and 0.31, respectively). There was no significant difference in the GATA1 mutation frequency between those classified as black, white, or Asian, but Hispanics had a borderline non-significant increase in frequency of GATA1 mutations compared to non-Hispanics (8.5% compared to 4.0%, p=0.06). Based on data from the New York Cancer Registry reviewed in 2005, two of the patients with a GATA1 mutation subsequently developed leukemia; one patient developed AMKL and the other had a leukemia of unspecified phenotype. Out of the 547 GATA1-negative patients, there was only one case of leukemia, but it was AML excluding AMKL. These results confirm a pre-natal origin of GATA1 mutations in DS patients. The frequency in this study was lower than the 10 percent previously reported (2 of 21 DS blood spots). Obtaining Gunthrie card blood spots for GATA1 mutation analysis serves as a relatively non-invasive screening approach. The presence of a GATA1 mutation at the time of birth might serve as a biomarker for an increased risk of developing DS-related AMKL.

Expression and Mutation Analysis of the Myc Antagonist Gene Mad1 in Acute Leukemia.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4331-4331
Author(s):  
Ling Pan ◽  
Xuejun Zhang ◽  
Xiaohui Suo ◽  
Fuxu Wang ◽  
Zhiyun Niu ◽  
...  
Keyword(s):  
Acute Leukemia ◽  
Point Mutation ◽  
Leucine Zipper ◽  
Mononuclear Cells ◽  
Single Strand Conformation Polymorphism ◽  
Polymorphism Analysis ◽  
Structural Domain ◽  
Significant Implication ◽  
Peripheral Blood Mononuclear

Abstract Objectives: Overexpression of Myc genes has been observed in many solid tumors and hematological malignancies, and Myc proteins are believed to act as transcription factors. In the past decade, several Myc-related genes have been identified, and the interaction mechanisms between these genes have been clarified. Members of Myc superfamily proteins belong to four different families: the Max family of common dimerization partners (including Max and MLx), the Myc family of proto-oncoproteins (including c-, N-, and L-Myc), the Myc antagonists including Mad family (including Mad1 and Mxi1) and Rox (Mnt). Mad1 gene maps to human chromosome 2p13 and encodes a 221-amino acid protein. Mad1 protein was shown to counteract the effects of Myc proteins and to inhibit the proliferation and tumorigenicity of glioblastoma and astrocytoma cells, and therefore was considered to be a kind of tumor suppressor. All Myc super-family members share a highly conserved structural domain, basic helix-loop-helix leucine zipper domain (bHLHZip). This bHLHzip domain is essential for these proteins to function during molecular switching from proliferation to differentiation. To function in vivo, Myc and Mad1 must heterodimerize with Max through the HLHZip motif. Then through the b (basic) motif, the heterodimers bind with CACGTG (E-box) containing DNA to activate or inactivate transcription. The present study is thus designed to examine the expression of Mad1 gene and to search for mutations in bHLHzip encoding domain. Methods: we screened bone marrow mononuclear cells (BMMNC) from 26 patients with AL and peripheral blood mononuclear cells (PBMNC) from 30 healthy volunteers, using reverse transcription- polymerase chain reaction, single strand conformation polymorphism analysis and sequencing. Results: Mad1 gene was expressed in all samples. Two polymorphisms of Mad1 were found in BMMNC from AL patients and PBMNC from health volunteers. A point mutation was detected at a very important position, a substitution of A397G inducing Lys78Glu in two patients. This might have significant implication for the function of Mad1, as Mad1 will heterodimerize with Max at this very point. Mutation at this point could thus disrupt proper intermolecular alignment between Mad1 and Max. The two patients with Mad1 gene mutation had poor clinical outcomes, patient No.9 never achieved CR and died 2 months after diagnosis, while patient No.11 got CR, after two courses chemotherapy, but relapsed quickly and died 3 months after diagnosis. Conclusion: Mad1 gene expressed and displayed mutations in acute leukemia patients. It would be interesting to know whether these mutations influence either the heterodimerizing or DNA binding potentials of Mad1 protein, by performing functional analysis of the mutant protein.

scholarly journals Association of DC-SIGN Promoter Polymorphism with Increased Risk for Parenteral, but Not Mucosal, Acquisition of Human Immunodeficiency Virus Type 1 Infection

2004 ◽  
Vol 78 (24) ◽  
pp. 14053-14056 ◽  
Author(s):  
Maureen P. Martin ◽  
Michael M. Lederman ◽  
Holli B. Hutcheson ◽  
James J. Goedert ◽  
George W. Nelson ◽  
...  
Keyword(s):  
At Risk ◽  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Single Strand Conformation Polymorphism ◽  
Nucleotide Polymorphisms ◽  
Increased Risk ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT There is considerable debate about the fundamental mechanisms that underlie and restrict acquisition of human immunodeficiency virus type 1 (HIV-1) infection. In light of recent studies demonstrating the ability of C type lectins to facilitate infection with HIV-1, we explored the potential relationship between polymorphisms in the DC-SIGN promoter and risk for acquisition of HIV-1 according to route of infection. Using samples obtained from 1,611 European-American participants at risk for parenteral (n = 713) or mucosal (n = 898) infection, we identified single-nucleotide polymorphisms in the DC-SIGN promoter using single-strand conformation polymorphism. Individuals at risk for parenterally acquired infection who had −336C were more susceptible to infection than were persons with −336T (odds ratio = 1.87, P = 0.001). This association was not observed in those at risk for mucosally acquired infection. A potential role for DC-SIGN specific to systemic acquisition and dissemination of infection is suggested.

scholarly journals Mutational activation of N- and K-ras oncogenes in plasma cell dyscrasias

Blood ◽  
1993 ◽  
Vol 81 (10) ◽  
pp. 2708-2713 ◽  
Author(s):  
P Corradini ◽  
M Ladetto ◽  
C Voena ◽  
A Palumbo ◽  
G Inghirami ◽  
...  
Keyword(s):  
Plasma Cell ◽  
Direct Sequencing ◽  
Single Strand Conformation Polymorphism ◽  
Polymorphism Analysis ◽  
Ras Mutations ◽  
Stage Disease ◽  
Plasma Cell Dyscrasias ◽  
Genomic Dnas ◽  
Multiple Myelomas ◽  
Mutational Activation

Abstract The frequency of N- and K-ras oncogene mutations was investigated in plasma cell dyscrasias. Genomic DNAs from 128 patients were selected for this study: 30 monoclonal gammopathies of undetermined significance, 8 solitary plasmacytomas, 77 multiple myelomas (MM), and 13 plasma cell leukemias (PCL). A two-step experimental approach was devised. All samples were screened for mutations by single-strand conformation polymorphism analysis. DNA fragments displaying an altered electrophoretic mobility were further studied by direct sequencing to confirm and characterize the nature of the mutations. Ras mutations are not randomly distributed because they are detectable only in MM (9%) and PCL (30.7%). N-ras codons 12, 13, and 61 and K-ras codon 12 were found to be mutated, but N-ras codon 61 mutation was the most frequent finding (63.6%). In conclusion, ras mutations were found in PCL, and in a subset of MM characterized by advanced-stage disease and adverse prognostic parameters. Furthermore, based on our findings, it is possible to speculate that ras mutations represent a late molecular lesion in the process of multistep carcinogenesis.

scholarly journals Molecular study of eight Japanese cases of glucose-6-phosphate dehydrogenase deficiency by nonradioisotopic single-strand conformation polymorphism analysis [see comments]

Blood ◽  
1994 ◽  
Vol 83 (11) ◽  
pp. 3363-3368 ◽  
Author(s):  
A Hirono ◽  
S Miwa ◽  
H Fujii ◽  
F Ishida ◽  
K Yamada ◽  
...  
Keyword(s):  
Direct Sequencing ◽  
Japanese Patients ◽  
Single Strand Conformation Polymorphism ◽  
Single Strand ◽  
Sscp Analysis ◽  
Polymorphism Analysis ◽  
Missense Mutations ◽  
G6pd Gene ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Conformation Polymorphism

Abstract Using a newly developed nonradioisotopic method of polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis combined with the direct sequencing using the fluorescence-labeled terminator, we identified seven missense mutations, 527 A-->G, 1003 G-- >A, 1159 C--eT, 1160 G-->A, 1229 G-->A, 1246 G-->A, and 1361 G-->A, in eight Japanese patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Except for the 527 A-->G, each mutation has been reported to cause variants G6PD Chatham, G6PD Guadalajara, G6PD Beverly Hills, G6PD “Japan”, G6PD Tokyo, and G6PD Andalus, respectively. In addition, a single base deletion in intron 5 was found in the patients with G6PD Guadalajara or G6PD Andalus. The variant with unique 527 A-->G was characterized and designated as G6PD Shinshu. We also characterized G6PD “Japan” and found that the variant had the striking resemblance with G6PD Riverside, bearing a missense mutation in the same codon, but causing a different amino acid substitution. Our modified PCR-SSCP analysis using minigel and ethidium bromide staining could detect six of the eight diverse mutations in the G6PD gene. Because it is easy and requires no special apparatus, this modified method will be useful for screening mutations in the G6PD gene.

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