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1.  Transient structure and dynamics in the disordered c-Myc transactivation domain affect Bin1 binding 
Nucleic Acids Research  2012;40(13):6353-6366.
The crucial role of Myc as an oncoprotein and as a key regulator of cell growth makes it essential to understand the molecular basis of Myc function. The N-terminal region of c-Myc coordinates a wealth of protein interactions involved in transformation, differentiation and apoptosis. We have characterized in detail the intrinsically disordered properties of Myc-1–88, where hierarchical phosphorylation of S62 and T58 regulates activation and destruction of the Myc protein. By nuclear magnetic resonance (NMR) chemical shift analysis, relaxation measurements and NOE analysis, we show that although Myc occupies a very heterogeneous conformational space, we find transiently structured regions in residues 22–33 and in the Myc homology box I (MBI; residues 45–65); both these regions are conserved in other members of the Myc family. Binding of Bin1 to Myc-1–88 as assayed by NMR and surface plasmon resonance (SPR) revealed primary binding to the S62 region in a dynamically disordered and multivalent complex, accompanied by population shifts leading to altered intramolecular conformational dynamics. These findings expand the increasingly recognized concept of intrinsically disordered regions mediating transient interactions to Myc, a key transcriptional regulator of major medical importance, and have important implications for further understanding its multifaceted role in gene regulation.
PMCID: PMC3401448  PMID: 22457068
2.  Psychometric validation of the Dutch translation of the quality of life in reflux and dyspepsia (QOLRAD) questionnaire in patients with gastroesophageal reflux disease 
The Quality of Life in Reflux and Dyspepsia (QOLRAD) questionnaire is one of the best-characterized disease-specific instruments that captures health-related problems and symptom-patterns in patients with gastroesophageal reflux disease (GERD). This paper reports the psychometric validation of a Dutch translation of the QOLRAD questionnaire in gastroenterology outpatients with GERD.
Patients completed the QOLRAD questionnaire at visit 1 (baseline), visit 2 (after 2, 4 or 8 weeks of acute treatment with esomeprazole 40 mg once daily), and visit 4 (after 6 months with on-demand esomeprazole 40 mg once daily or continuous esomeprazole 20 mg once daily). Symptoms were assessed at each visit, and patient satisfaction was assessed at visits 2 and 4.
Of the 1166 patients entered in the study, 97.3% had moderate or severe heartburn and 55.5% had moderate or severe regurgitation at baseline. At visit 2, symptoms of heartburn and regurgitation were mild or absent in 96.7% and 97.7%, respectively, and 95.3% of patients reported being satisfied with the treatment. The internal consistency and reliability of the QOLRAD questionnaire (range: 0.83-0.92) supported construct validity. Convergent validity was moderate to low. Known-groups validity was confirmed by a negative correlation between the QOLRAD score and clinician-assessed severity of GERD symptoms. Effect sizes (1.15-1.93) and standardized response means (1.17-1.86) showed good responsiveness to change. GERD symptoms had a negative impact on patients' lives.
The psychometric characteristics of the Dutch translation of the QOLRAD questionnaire were found to be satisfactory, with good reliability and responsiveness to change, although convergent validity was at best moderate.
PMCID: PMC2933707  PMID: 20716328
3.  Validity and reliability of a new, short symptom rating scale in patients with persistent atrial fibrillation 
Symptoms related to atrial fibrillation and their impact on health-related quality of life (HRQoL) are often evaluated in clinical trials. However, there remains a need for a properly validated instrument. We aimed to develop and validate a short symptoms scale for patients with AF.
One hundred and eleven patients with a variety of symptoms related to AF were scheduled for DC cardioversion. The mean age was 67.1 ± 12.1 years, and 80% were men. The patients completed the new symptoms scale, the Toronto Symptoms Check List (SCL) and the generic Short Form 36 (SF-36) the day before the planned DC cardioversion. Compliance was excellent, with only 1 of 666 answers missing.
One item, 'limitations in working capability', was deleted because of a low numerical response rate, as many of the patients were retired. The internal consistency reliability of the remaining six items was 0.81 (Cronbach's α). Patients scored highest in the items of 'dyspnoea on exertion', 'limitations in daily life due to AF' and 'fatigue due to AF', with scores of 4.5, 3.3 and 4.5, respectively. There was a good correlation to all relevant SF-36 domains and to the relevant questions of the SCL. The Rasch analyses showed that the items are unidimensional and that they are clearly separated and cover an adequate range. Test-retest reliability was performed in patients who failed DC and was adequate for three of six items, >0.70.
The psychometric characteristics of the new short symptoms scale were found to have satisfactory reliability and validity.
PMCID: PMC2717073  PMID: 19604399
4.  Reliability and validity of the Gastrointestinal Symptom Rating Scale (GSRS) and Quality of Life in Reflux and Dyspepsia (QOLRAD) questionnaire in dyspepsia: A six-country study 
Symptoms of dyspepsia significantly disrupt patients' lives and reliable methods of assessing symptom status are important for patient management. The aim of the current study was to document the psychometric characteristics of the Gastrointestinal Symptom Rating Scale (GSRS) and the Quality of Life in Reflux and Dyspepsia questionnaire (QOLRAD) in Afrikaans, German, Hungarian, Italian, Polish and Spanish patients with dyspepsia.
853 patients with symptoms of dyspepsia completed the GSRS, the QOLRAD, the 36-item Short-Form Health Survey (SF-36) and the Hospital Anxiety and Depression scale.
The internal consistency reliability of the GSRS was 0.43–0.87 and of the QOLRAD 0.79–0.95. Test-retest reliability of the GSRS was 0.36–0.75 and of the QOLRAD 0.41–0.82. GSRS Abdominal pain domain correlated significantly with all QOLRAD domains in most language versions, and with SF-36 Bodily pain in all versions. QOLRAD domains correlated significantly with the majority of SF-36 domains in most versions. Both questionnaires were able to differentiate between patients whose health status differed according to symptom frequency and severity.
The psychometric characteristics of the different language versions of the GSRS and QOLRAD were found to be good, with acceptable reliability and validity. The GSRS and QOLRAD were found to be useful for evaluating dyspeptic symptoms and their impact on patients' daily lives in multinational clinical trials.
PMCID: PMC2276197  PMID: 18237386
5.  An instrument for measuring health-related quality of life in patients with Deep Venous Thrombosis (DVT): development and validation of Deep Venous Thrombosis Quality of Life (DVTQOL) questionnaire 
Few studies have evaluated patient-reported outcomes in connection with a primary event of deep venous thrombosis, partly due to a lack of disease-specific measures. The aim here was to develop a disease-specific health-related quality of life (HRQL) measure, the deep venous thrombosis quality of life questionnaire (DVTQOL), for patients with recent exposition and treatment of proximal deep venous thrombosis.
A total of 121 consecutive outpatients (50 % males; mean age 61.2 ± 14 years) treated with warfarin (Waran®) for symptomatic proximal deep venous thrombosis were included in the study. Patients completed the SF-36, EQ-5D and the pilot version of the DVTQOL.
Items having: high ceiling and floor effect, items with lower factor loadings than 0.50 and items loading in several factors were removed from the pilot version of DVTQOL. In addition, overlapping and redundant items identified by the Rasch analysis were excluded. The final DVTQOL questionnaire consists of 29 items composing six dimensions depicting problems with: emotional distress; symptoms (e.g. pain, swollen ankles, cramp, bruising); limitation in physical activity; hassle with coagulation monitoring; sleep disturbance; and dietary problems. The internal consistency reliability was high (alpha value ranged from 0.79 to 0.93). The relevant domains of the SF-36 and EQ-5D significantly correlated with DVTQOL, thereby confirming its construct validity.
The DVTQOL is a short and user-friendly instrument with good reliability and validity. Its test-retest reliability and responsiveness to change in clinical trials, however, must be explored.
PMCID: PMC471562  PMID: 15214965
Deep Venous Thromboses; Health-related Quality of Life; Psychometric evaluation
6.  Psychometric validation of the German translation of the Gastrointestinal Symptom Rating Scale (GSRS) and Quality of Life in Reflux and Dyspepsia (QOLRAD) questionnaire in patients with reflux disease 
Symptoms of heartburn has an impact on health-related quality of life (HRQL). When a questionnaire is translated into a new language, a linguistic validation is necessary but not sufficient unless the psychometric characteristics have been verified. The aim is to document the psychometric characteristics of the German translation of the Gastrointestinal Symptom Rating Scale (GSRS) and Quality of Life in Reflux and Dyspepsia (QOLRAD) questionnaire.
142 patients with symptoms of heartburn (Age: M = 47.5, ± 14.6; Males = 44.4%) completed the German translation of GSRS, the QOLRAD, the Short-Form-36 (SF-36) and the Hospital Anxiety and Depression (HAD) scale.
The internal consistency reliability of GSRS ranged from 0.53–0.91 and of QOLRAD from 0.90–0.94, respectively. The test-retest reliability of GSRS ranged from 0.49–0.73 and of QOLRAD from 0.70–0.84. The relevant domains of the GSRS and QOLRAD domain scores significantly correlated. GSRS domains of Abdominal Pain and Constipation correlated (negatively) with most of the domains of the SF-36. The relevant QOLRAD domains significantly correlated with all SF-36 domains.
The psychometric characteristics of the German translation of GSRS and QOLRAD were found to be good, with satisfactory reliability and validity. The reliability of the GSRS Abdominal Pain domain was moderate.
PMCID: PMC269999  PMID: 14613560
Reliability; Validity; Reflux; Health-Related Quality of Life; Germany
7.  Investigation and prediction of the severity of p53 mutants using parameters from structural calculations 
The Febs Journal  2009;276(15):4142-4155.
A method has been developed to predict the effects of mutations in the p53 cancer suppressor gene. The new method uses novel parameters combined with previously established parameters. The most important parameter is the stability measure of the mutated structure calculated using molecular modelling. For each mutant, a severity score is reported, which can be used for classification into deleterious and nondeleterious. Both structural features and sequence properties are taken into account. The method has a prediction accuracy of 77% on all mutants and 88% on breast cancer mutations affecting WAF1 promoter binding. When compared with earlier methods, using the same dataset, our method clearly performs better. As a result of the severity score calculated for every mutant, valuable knowledge can be gained regarding p53, a protein that is believed to be involved in over 50% of all human cancers.
PMCID: PMC2730554  PMID: 19558493
cancer; molecular modelling; mutations; p53; structural prediction
8.  Functionally Important Amino Acids in the Arabidopsis Thylakoid Phosphate Transporter: Homology Modeling and Site-Directed Mutagenesis† 
Biochemistry  2010;49(30):6430-6439.
The anion transporter 1 (ANTR1) from Arabidopsis thaliana, homologous to the mammalian members of the solute carrier 17 (SLC17) family, is located in the chloroplast thylakoid membrane. When expressed heterologously in Escherichia coli, ANTR1 mediates a Na+-dependent active transport of inorganic phosphate (Pi). The aim of this study was to identify amino acid residues involved in Pi binding and translocation by ANTR1 and in the Na+ dependence of its activity. A three-dimensional structural model of ANTR1 was constructed using the crystal structure of glycerol 3-phosphate/phosphate antiporter from E. coli as a template. Based on this model and multiple sequence alignments, five highly conserved residues in plant ANTRs and mammalian SLC17 homologues have been selected for site-directed mutagenesis, namely, Arg-120, Ser-124, and Arg-201 inside the putative translocation pathway and Arg-228 and Asp-382 exposed at the cytoplasmic surface of the protein. The activities of the wild-type and mutant proteins have been analyzed using expression in E. coli and radioactive Pi transport assays and compared with bacterial cells carrying an empty plasmid. The results from Pi- and Na+-dependent kinetics indicate the following: (i) Arg-120 and Arg-201 may be important for binding and translocation of the substrate; (ii) Ser-124 may function as a transient binding site for Na+ ions in close proximity to the periplasmic side; (iii) Arg-228 and Asp-382 may participate in interactions associated with protein conformational changes required for full transport activity. Functional characterization of ANTR1 should provide useful insights into the function of other plant and mammalian SLC17 homologous transporters.
PMCID: PMC2911078  PMID: 20565143

Results 1-8 (8)