11 results on '"Brault K"'
Search Results
2. Absent or shortened nasal bone length and the detection of Down Syndrome in second-trimester fetuses.
- Author
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Gianferrari EA, Benn PA, Dries L, Brault K, Egan JFX, Zelop CM, Gianferrari, Elisa A, Benn, Peter A, Dries, Lisa, Brault, Kim, Egan, James F X, and Zelop, Carolyn M
- Published
- 2007
- Full Text
- View/download PDF
3. Improving biophysical properties of a bispecific antibody scaffold to aid developability: quality by molecular design.
- Author
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Von Kreudenstein TS, Escobar-Carbrera E, Lario PI, D'Angelo I, Brault K, Kelly J, Durocher Y, Baardsnes J, Woods RJ, Xie MH, Girod PA, Suits MD, Boulanger MJ, Poon DK, Ng GY, and Dixit SB
- Subjects
- Animals, Antibodies chemistry, Antibodies genetics, Antibodies, Bispecific chemistry, Antibodies, Bispecific genetics, CHO Cells, Chromatography, Liquid, Cricetinae, Cricetulus, Drug Design, Humans, Hydrogen-Ion Concentration, Hydrophobic and Hydrophilic Interactions, Immunoglobulin Fc Fragments chemistry, Immunoglobulin Fc Fragments genetics, Mass Spectrometry, Models, Molecular, Mutation, Protein Engineering methods, Protein Multimerization, Protein Structure, Tertiary, Static Electricity, Temperature, Antibodies immunology, Antibodies, Bispecific immunology, Antibody Specificity immunology, Immunoglobulin Fc Fragments immunology
- Abstract
While the concept of Quality-by-Design is addressed at the upstream and downstream process development stages, we questioned whether there are advantages to addressing the issues of biologics quality early in the design of the molecule based on fundamental biophysical characterization, and thereby reduce complexities in the product development stages. Although limited number of bispecific therapeutics are in clinic, these developments have been plagued with difficulty in producing materials of sufficient quality and quantity for both preclinical and clinical studies. The engineered heterodimeric Fc is an industry-wide favorite scaffold for the design of bispecific protein therapeutics because of its structural, and potentially pharmacokinetic, similarity to the natural antibody. Development of molecules based on this concept, however, is challenged by the presence of potential homodimer contamination and stability loss relative to the natural Fc. We engineered a heterodimeric Fc with high heterodimeric specificity that also retains natural Fc-like biophysical properties, and demonstrate here that use of engineered Fc domains that mirror the natural system translates into an efficient and robust upstream stable cell line selection process as a first step toward a more developable therapeutic.
- Published
- 2013
- Full Text
- View/download PDF
4. Characterization of papillomavirus E1 helicase mutants defective for interaction with the SUMO-conjugating enzyme Ubc9.
- Author
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Fradet-Turcotte A, Brault K, Titolo S, Howley PM, and Archambault J
- Subjects
- Amino Acid Sequence, Amino Acid Substitution, DNA-Binding Proteins genetics, Gene Expression Regulation physiology, Human papillomavirus 11 genetics, Humans, Models, Molecular, Molecular Sequence Data, Mutation, Organisms, Genetically Modified, Protein Conformation, Protein Transport physiology, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae metabolism, Ubiquitin-Conjugating Enzymes genetics, Viral Proteins genetics, DNA-Binding Proteins metabolism, Human papillomavirus 11 enzymology, Small Ubiquitin-Related Modifier Proteins metabolism, Ubiquitin-Conjugating Enzymes metabolism, Viral Proteins metabolism
- Abstract
The E1 helicase from BPV and HPV16 interacts with Ubc9 to facilitate viral genome replication. We report that HPV11 E1 also interacts with Ubc9 in vitro and in the yeast two-hybrid system. Residues in E1 involved in oligomerization (353-435) were sufficient for binding to Ubc9 in vitro, but the origin-binding and ATPase domains were additionally required in yeast. Nuclear accumulation of BPV E1 was shown previously to depend on its interaction with Ubc9 and sumoylation on lysine 514. In contrast, HPV11 and HPV16 E1 mutants defective for Ubc9 binding remained nuclear even when the SUMO pathway was inhibited. Furthermore, we found that K514 in BPV E1 and the analogous K559 in HPV11 E1 are not essential for nuclear accumulation of E1. These results suggest that the interaction of E1 with Ubc9 is not essential for its nuclear accumulation but, rather, depends on its oligomerization and binding to DNA and ATP.
- Published
- 2009
- Full Text
- View/download PDF
5. Variation of fetal nasal bone length in second-trimester fetuses according to race and ethnicity.
- Author
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Zelop CM, Milewski E, Brault K, Benn P, Borgida AF, and Egan JF
- Subjects
- Female, Gestational Age, Humans, Pregnancy, Retrospective Studies, Black or African American, Asian, Hispanic or Latino, Nasal Bone diagnostic imaging, Nasal Bone embryology, Ultrasonography, Prenatal, White People
- Abstract
Objective: The purpose of this study was to determine the influence of race and ethnicity on the expected nasal bone length (NBL) based on biparietal diameter (BPD) measured in second-trimester fetuses., Methods: We searched our ultrasound, obstetric, and cytogenetic databases for all second-trimester fetuses with measured NBLs. Fetuses with Down syndrome were identified and excluded from the analysis. Linear regression curves were generated for NBL by BPD according to race and ethnicity. Categories used were African American, Hispanic, Asian, and white. Analysis of variance was used to compare mean variation of observed from expected NBL by BPD according to race and ethnicity., Results: There were 717 fetuses with NBL-by-BPD pairs who were available for analysis in our population, including 139 African American, 58 Hispanic, 22 Asian, and 498 white fetuses. Nasal bone length was highly correlated with BPD for each race (P < .001). Mean variances of observed from expected NBL by BPD were statistically different according to race or ethnicity (P = .0092)., Conclusions: Race and ethnicity significantly affect the mean regression line of expected NBL by BPD among fetuses in the second trimester. Genetic sonographic norms, therefore, appear to require race- and ethnicity-specific formulas for NBL.
- Published
- 2005
- Full Text
- View/download PDF
6. Inhibition of human papillomavirus DNA replication by small molecule antagonists of the E1-E2 protein interaction.
- Author
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White PW, Titolo S, Brault K, Thauvette L, Pelletier A, Welchner E, Bourgon L, Doyon L, Ogilvie WW, Yoakim C, Cordingley MG, and Archambault J
- Subjects
- Amino Acid Substitution, Animals, Base Sequence, Binding Sites, CHO Cells, Cottontail rabbit papillomavirus drug effects, Cottontail rabbit papillomavirus genetics, Cottontail rabbit papillomavirus metabolism, Cricetinae, DNA, Viral genetics, DNA-Binding Proteins antagonists & inhibitors, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Humans, Oncogene Proteins, Viral antagonists & inhibitors, Oncogene Proteins, Viral genetics, Oncogene Proteins, Viral metabolism, Papillomaviridae genetics, Protein Binding drug effects, Rabbits, Recombinant Proteins antagonists & inhibitors, Recombinant Proteins genetics, Recombinant Proteins metabolism, Transcriptional Activation, Transfection, Viral Proteins genetics, DNA Replication drug effects, DNA, Viral biosynthesis, Papillomaviridae drug effects, Papillomaviridae metabolism, Viral Proteins antagonists & inhibitors, Viral Proteins metabolism
- Abstract
Human papillomavirus (HPV) DNA replication is initiated by recruitment of the E1 helicase by the E2 protein to the viral origin. Screening of our corporate compound collection with an assay measuring the cooperative binding of E1 and E2 to the origin identified a class of small molecule inhibitors of the protein interaction between E1 and E2. Isothermal titration calorimetry and changes in protein fluorescence showed that the inhibitors bind to the transactivation domain of E2, the region that interacts with E1. These compounds inhibit E2 of the low risk HPV types 6 and 11 but not those of high risk HPV types or of cottontail rabbit papillomavirus. Functional evidence that the transactivation domain is the target of inhibition was obtained by swapping this domain between a sensitive (HPV11) and a resistant (cottontail rabbit papillomavirus) E2 type and by identifying an amino acid substitution, E100A, that increases inhibition by approximately 10-fold. This class of inhibitors was found to antagonize specifically the E1-E2 interaction in vivo and to inhibit HPV DNA replication in transiently transfected cells. These results highlight the potential of the E1-E2 interaction as a small molecule antiviral target.
- Published
- 2003
- Full Text
- View/download PDF
7. Characterization of recombinant HPV6 and 11 E1 helicases: effect of ATP on the interaction of E1 with E2 and mapping of a minimal helicase domain.
- Author
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White PW, Pelletier A, Brault K, Titolo S, Welchner E, Thauvette L, Fazekas M, Cordingley MG, and Archambault J
- Subjects
- Adenosine Triphosphatases chemistry, Adenosine Triphosphatases metabolism, Amino Acid Sequence, Amino Acid Substitution, Base Sequence, Biopolymers, Catalysis, DNA Helicases chemistry, DNA Primers, Molecular Sequence Data, Papillomaviridae isolation & purification, Protein Folding, Protein Structure, Quaternary, Recombinant Proteins chemistry, Recombinant Proteins metabolism, Substrate Specificity, Adenosine Triphosphate metabolism, DNA Helicases metabolism, Papillomaviridae enzymology
- Abstract
To better characterize the enzymatic activities required for human papillomavirus (HPV) DNA replication, the E1 helicases of HPV types 6 and 11 were produced using a baculovirus expression system. The purified wild type proteins and a version of HPV11 E1 lacking the N-terminal 71 amino acids, which was better expressed, were found to be hexameric over a wide range of concentrations and to have helicase and ATPase activities with relatively low values for K(m)(ATP) of 12 microm for HPV6 E1 and 6 microm for HPV11 E1. Interestingly, the value of K(m)(ATP) was increased 7-fold in the presence of the E2 transactivation domain. In turn, ATP was found to perturb the co-operative binding of E1 and E2 to DNA. Mutant and truncated versions of in vitro translated E1 were used to identify a minimal ATPase domain composed of the C-terminal 297 amino acids. This fragment was expressed, purified, and found to be fully active in ATP hydrolysis, single-stranded DNA binding, and unwinding assays, despite lacking the minimal origin-binding domain.
- Published
- 2001
- Full Text
- View/download PDF
8. Aromatic hydrocarbon receptor (AhR).AhR nuclear translocator- and p53-mediated induction of the murine multidrug resistance mdr1 gene by 3-methylcholanthrene and benzo(a)pyrene in hepatoma cells.
- Author
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Mathieu MC, Lapierre I, Brault K, and Raymond M
- Subjects
- Animals, Aryl Hydrocarbon Receptor Nuclear Translocator, Benzo(a)pyrene pharmacology, Carcinoma, Hepatocellular, Mice, Models, Biological, Mutation, Promoter Regions, Genetic, RNA, Messenger biosynthesis, Receptors, Aryl Hydrocarbon genetics, Response Elements, Transcription Factors biosynthesis, Transcription Factors genetics, Transcriptional Activation, Tumor Cells, Cultured, Tumor Suppressor Protein p53 biosynthesis, Xenobiotics pharmacology, ATP Binding Cassette Transporter, Subfamily B, Member 1 biosynthesis, DNA-Binding Proteins, Genes, MDR, Methylcholanthrene pharmacology, Receptors, Aryl Hydrocarbon physiology, Transcription Factors physiology, Tumor Suppressor Protein p53 physiology
- Abstract
The mouse multidrug resistance gene family consists of three genes (mdr1, mdr2, and mdr3) encoding P-glycoprotein. We show that the expression of mdr1 is increased at the transcriptional level upon treatment of the hepatoma cell line Hepa-1c1c7 with the polycyclic aromatic hydrocarbon 3-methylcholanthrene (3-MC). This increase is not observed in the aromatic hydrocarbon receptor (AhR)-defective TAOc1BP(r)c1 and the AhR nuclear translocator (Arnt)-defective BP(r)c1 variants, demonstrating that the induction of mdr1 by 3-MC requires AhR.Arnt. We show that the mdr1 promoter (-1165 to +84) is able to activate the expression of a reporter gene in response to 3-MC in Hepa-1c1c7 but not in BP(r)c1 cells. Deletion analysis indicated that the region from -245 to -141 contains cis-acting sequences mediating the induction, including a potential p53 binding sequence. 3-MC treatment of the cells increased the levels of p53 and induced p53 binding to the mdr1 promoter in an AhR.Arnt-dependent manner. Mutations in the p53 binding site abrogated induction of mdr1 by 3-MC, indicating that p53 binding to the mdr1 promoter is essential for the induction. Benzo(a)pyrene, a polycyclic aromatic hydrocarbon and AhR ligand, which, like 3-MC, is oxidized by metabolizing enzymes regulated by AhR.Arnt, also activated p53 and induced mdr1 transcription. 2,3,7,8-Tetrachlorodibenzo-p-dioxin, an AhR ligand resistant to metabolic breakdown, had no effect. These results indicate that the transcriptional induction of mdr1 by 3-MC and benzo(a)pyrene is directly mediated by p53 but that the metabolic activation of these compounds into reactive species is necessary to trigger p53 activation. The ability of the anticancer drug and potent genotoxic agent daunorubicin to induce mdr1 independently of AhR.Arnt further supports the proposition that mdr1 is transcriptionally up-regulated by p53 in response to DNA damage.
- Published
- 2001
- Full Text
- View/download PDF
9. Identification of domains of the human papillomavirus type 11 E1 helicase involved in oligomerization and binding to the viral origin.
- Author
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Titolo S, Pelletier A, Pulichino AM, Brault K, Wardrop E, White PW, Cordingley MG, and Archambault J
- Subjects
- Adenosine Triphosphatases metabolism, Adenosine Triphosphate metabolism, Amino Acid Sequence, Amino Acid Substitution, Conserved Sequence, DNA Replication, DNA, Single-Stranded metabolism, DNA-Binding Proteins genetics, Humans, Papillomaviridae genetics, Papillomaviridae metabolism, Protein Structure, Tertiary, Two-Hybrid System Techniques, Viral Proteins genetics, DNA, Viral metabolism, DNA-Binding Proteins chemistry, DNA-Binding Proteins metabolism, Papillomaviridae chemistry, Replication Origin, Viral Proteins chemistry, Viral Proteins metabolism
- Abstract
The E1 helicase of papillomavirus is required, in addition to host cell DNA replication factors, during the initiation and elongation phases of viral episome replication. During initiation, the viral E2 protein promotes the assembly of enzymatically active multimeric E1 complexes at the viral origin of DNA replication. In this study we used the two-hybrid system and chemical cross-linking to demonstrate that human papillomavirus type 11 (HPV11) E1 can self-associate in yeast and form hexamers in vitro in a reaction stimulated by single-stranded DNA. Self-association in yeast was most readily detected using constructs spanning the E1 C-terminal domain (amino acids 353 to 649) and was dependent on a minimal E1-E1 interaction region located between amino acids 353 and 431. The E1 C-terminal domain was also able to oligomerize in vitro but, in contrast to wild-type E1, did so efficiently in the absence of single-stranded DNA. Sequences located between amino acids 191 and 353 were necessary for single-stranded DNA to modulate oligomerization of E1 and were also required, together with the rest of the C terminus, for binding of E1 to the origin. Two regions within the C-terminal domain were identified as important for oligomerization: the ATP-binding domain and region A, which is located within the minimal E1-E1 interaction domain and is one of four regions of E1 that is highly conserved with the large T antigens of simian virus 40 and polyomavirus. Amino acid substitutions of highly conserved residues within the ATP-binding domain and region A were identified that reduced the ability of E1 to oligomerize and bind to the origin in vitro and to support transient DNA replication in vivo. These results support the notion that oligomerization of E1 occurs primarily through the C-terminal domain of the protein and is allosterically regulated by DNA and ATP. The bipartite organization of the E1 C-terminal domain is reminiscent of that found in other hexameric proteins and suggests that these proteins may oligomerize by a similar mechanism.
- Published
- 2000
- Full Text
- View/download PDF
10. Role of the ATP-binding domain of the human papillomavirus type 11 E1 helicase in E2-dependent binding to the origin.
- Author
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Titolo S, Pelletier A, Sauvé F, Brault K, Wardrop E, White PW, Amin A, Cordingley MG, and Archambault J
- Subjects
- Amino Acid Substitution, Animals, Binding Sites, Cattle, DNA Helicases genetics, DNA Replication, DNA-Binding Proteins genetics, Glutamic Acid genetics, Glutamic Acid metabolism, Glutamine genetics, Glutamine metabolism, Humans, Lysine genetics, Lysine metabolism, Magnesium, Papillomaviridae genetics, Papillomaviridae physiology, Proline genetics, Proline metabolism, Saccharomyces cerevisiae, Serine genetics, Serine metabolism, Temperature, Viral Proteins genetics, Virus Replication, Adenosine Triphosphate metabolism, DNA Helicases metabolism, DNA-Binding Proteins metabolism, Papillomaviridae metabolism, Replication Origin, Viral Proteins metabolism
- Abstract
Replication of the genome of human papillomaviruses (HPV) is initiated by the recruitment of the viral E1 helicase to the origin of DNA replication by the viral E2 protein, which binds specifically to the origin. We determined, for HPV type 11 (HPV-11), that the C-terminal 296 amino acids of E1 are sufficient for interaction with the transactivation domain of E2 in the yeast two-hybrid system and in vitro. This region of E1 encompasses the ATP-binding domain. Here we have examined the role of this ATP-binding domain, and of ATP, on E2-dependent binding of E1 to the origin. Several amino acid substitutions in the phosphate-binding loop (P loop), which is implicated in binding the triphosphate moiety of ATP, abolished E2 binding, indicating that the structural integrity of this domain is essential for the interaction. The structural constraints imposed on the E1 P loop may differ between HPV-11 and bovine papillomavirus type 1 (BPV-1), since the P479S substitution that inactivates BPV-1 E1 is tolerated in the HPV-11 enzyme. Other substitutions in the E1 P loop, or in two other conserved motifs of the ATP-binding domain, were tolerated, indicating that ATP binding is not essential for interaction with E2. Nevertheless, ATP-Mg stimulated the E2-dependent binding of E1 to the origin in vitro. This stimulation was maximal at the physiological temperature (37 degrees C) and did not require ATP hydrolysis. In contrast, ATP-Mg did not stimulate the E2-dependent binding to the origin of an E1 protein containing only the C-terminal domain (353 to 649) or that of mutant E1 proteins with alterations in the DNA-binding domain. These results are discussed in light of a model in which the E1 ATP-binding domain is required for formation of the E2-binding surface and can, upon the binding of ATP, facilitate and/or stabilize the interaction of E1 with the origin.
- Published
- 1999
- Full Text
- View/download PDF
11. Maternal smoking and accelerated placental maturation.
- Author
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Pinette MG, Loftus-Brault K, Nardi DA, and Rodis JF
- Subjects
- Female, Gestational Age, Humans, Placenta pathology, Prospective Studies, Ultrasonography, Placenta physiology, Smoking adverse effects
- Abstract
In a prospective study of 367 patients referred for obstetric ultrasound evaluation, maternal cigarette smoking was found to have an important effect on the rate of placental maturation. The frequencies of grades 0, I, II, and III placentas throughout gestation were determined for both smoking and nonsmoking groups. Overall, the smoking mothers had more mature placentas than the nonsmoking mothers. This was reflected by the earlier mean gestational age at appearance of each of the placental grades in the smoking group as compared with the nonsmoking group. The mean gestational ages at appearance of grades 0, I, II, and III placentas for smoking mothers were 23.5, 27.9, 32, and 34.4 weeks, respectively, as compared with 26, 31.6, 35.7, and 38.3 weeks in the nonsmoking group. Smokers had a greater frequency of grade II placentas from 22-35 weeks than nonsmokers, and a greater frequency of grade III placentas beyond 25 weeks.
- Published
- 1989
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