Exploration of helicase mechanisms at single molecule level: answering decades old questions and bioengineering super proteins

In the last 15 years, single molecule techniques have allowed us to observe the transient, heterogeneous, and multi‐step behavior of biomolecules often averaged out and lost in ensemble assays. Helicases form a ubiquitous class of enzymes that function in many aspects of nucleic acid metabolism which is central to life. We used single molecule Förster resonance energy transfer (FRET) and optical tweezers force spectroscopy to study translocation and unwinding mechanism of Escherichia coli Rep, a model helicase with 3’5’ motor translocation activity on single‐stranded DNA (ssDNA) fueled by ATP hydrolysis. When a translocating Rep on a 3’‐overhang DNA reaches the DNA duplex junction, instead of dissociating, it snaps back to the 3’ end, restarting another shuttling cycle. We investigated the mechanism of repetitive shuttling and discovered the same behavior on various DNA substrates. We concluded that the repetitions are induced by the lack of ssDNA track ahead of the Rep. Using the repetitive shuttling assay, we explored the translocation mechanism of Rep in detail. First, we perturbed the ssDNA binding network via biochemical mutations that led to determination of the key residues that control shuttling speed, ATPase activity, and directionality. Second, we tested the effects of the DNA lesions on Rep translocation, observing that the irregularities encompassing 1‐3 nucleotides in the backbone and nucleobases only caused transient stalls. To probe the unwinding mechanism, we developed a conformational control assay which turned wild type Rep monomers with no detectable DNA unwinding activity into super‐helicases (Rep‐X) via internal covalent crosslinking. Rep‐X can unwind thousands of base pairs processively even against large forces. We also showed that partner proteins of a similar enzyme turn on the unwinding activity by stabilizing the active form. Lastly, the orientation dependence of FRET is experimentally shown between nucleic acid conjugated cyanine fluorophores that constitutes the first demonstration of this effect in a biophysical system since its formulation by Theodor Förster in 1948

The Validation of Solution Building Inventory in the Turkish Population

تعیین اعتبار فهرست‌سازی راه‌حل‌ها در جمعیت ترک زبان این مطالعه با هدف تطبیق فهرست‌سازی راه‌حل در ترک زبانان و بررسی ویژگی‌های روان‌سنجی آن انجام شده است. گروه مطالعه شده شامل 278 نفر (195 زن و 83 مرد) با میانگین سنی 18-23 سال بود. فهرست‌سازی راه‌حل ، مقیاس امید ذاتی و مقیاس تاثیر مثبت و منفی به عنوان ابزارهای اندازه‌گیری استفاده شدند. نتایج تحلیل عامل اکتشافی، تناسب داده‌ها با تحلیل عوامل و ساختار دو عامله را نشان داد که 49.38% از واریانس کل می‌باشد. یافته‌های تحلیل‌ عامل تاکیدی نشان داد که این مدل با ساختار دو بعدی 11 آیتمی این مقیاس، تناسب خوبی دارد. علاوه بر این، انتظار می‌رود ضرایب همبستگی روابط بین فهرست‌سازی راه‌حل و دیگر مقیاس‌ها، اعتباری معادل و همگرا ارائه دهد. در تحلیل‌های قابل اعتماد، ضریب هماهنگی داخلی آلفای کرونباخ برای این مقیاس 0.85 در مقیاس کل بود. در حالی که برای سیستم‌های حمایتی فرعی 0.78 و برای سیستم ایجاد شده فرعی 0.85 محاسبه شد. در نتیجه، فهرست‌سازی راه‌حل بعنوان ابزار اندازه‌گیری برای اثبات اعتبار و قابلیت اطمینان در ترک زبانان است

MAGiC: A multimodal framework for analysing gaze in dyadic communication

The analysis of dynamic scenes has been a challenging domain in eye tracking research. This study presents a framework, named MAGiC, for analyzing gaze contact and gaze aversion in face-to-face communication. MAGiC provides an environment that is able to detect and track the conversation partner’s face automatically, overlay gaze data on top of the face video, and incorporate speech by means of speech-act annotation. Specifically, MAGiC integrates eye tracking data for gaze, audio data for speech segmentation, and video data for face tracking. MAGiC is an open source framework and its usage is demonstrated via publicly available video content and wiki pages. We explored the capabilities of MAGiC through a pilot study and showed that it facilitates the analysis of dynamic gaze data by reducing the annotation effort and the time spent for manual analysis of video data

The Validation of Solution Building Inventory in the Turkish Population

This study aims to adapt Solution Building Inventory (SBI) to Turkish and examine its psychometric properties. The study group consisted of 278 individuals (195 females and 83 males), whose age range was between 18 and 23 years old. Solution Building Inventory, Dispositional Hope Scale, and Positive and Negative Affect Scale were used as measurements. The results of the exploratory factor analysis showed the suitability of the data for factor analysis and revealed a two-factor structure explaining 49.38% of the total variance. Confirmatory factor analysis findings showed that the model had a good fit in the 11-item and two-dimensional structure of the scale. Besides, the correlation coefficients of the relationship between the SBI and other scales were expectedly provided equivalent and convergent validity. In reliability analyzes, the Cronbach alpha internal consistency coefficient of the scale was found to be .85 for the scale in general, while it was calculated as .78 for the supporting solutions sub-dimension and .85 for the creating solutions sub-dimension. As a result, SBI is a measurement tool with proven validity and reliability in Turkish

Fracture resistance of roots filled with three different obturation techniques

Objectives: The aim of this study was to compare in vitro root fracture resistance following root canal filling with AH 26 using lateral condensation, BeeFill, and Thermafil techniques. Study Design: Eighty extracted human mandibular premolars with similar dimensions were selected. In order to standardize the roots, measurements were taken in two separate regions of the teeth?at the cemento-enamel junction and 8 mm apically from the junction?buccolingual as well as mesiodistal for every tooth. Teeth were then randomly divided into five groups (n=16). With the exception of the non-prepared group (Group 1), instrumentation was done in all groups. In group 2, instrumentation but no filling was performed; in group 3, the obturation was done with AH 26 + gutta-percha; in group 4, with AH 26 + BeeFill and in group 5, AH 26 + a Thermafil obturator was used. All the roots were mounted vertically in copper rings and filled with acrylic resin, exposing 8 mm of the coronal part. A universal testing machine was used for the strength test. Results: The results were analyzed using the one-way ANOVA test. The significance between the groups was tested with Temhane?s T2 test. The results indicate that instrumentation of root canals had a significant effect on fracture resistance (p0.05). Conclusions: The results suggest that instrumentation of root canals significantly weakens the tooth structure to fracture and the root canal obturation techniques that are used are not able to form reinforcement

Corticosteroid hypersensitivity in allergic rhinitis

Corticosteroid hypersensitivity in allergic rhinitis. Background: intranasal corticosteroid (IC) is the most effective treatment method in allergic rhinitis patients who are unresponsive to antihistamines. The literature reports an approximate 20% treatment failure for instances where IC is used for the treatment allergic rhinitis. Hypersensitivity reaction to corticosteroids may be one of the causes of this treatment failure. Objective: to discover the incidence and confounding factors of corticosteroid hypersensitivity in patients with allergic rhinitis. Methods: after 31 patients were excluded, 150 consecutive patients who were prospectively evaluated in our outpatient clinics with the diagnosis of allergic rhinitis and 50 age- and sex- matched healthy volunteers were included in this study. To diagnose allergic rhinitis, the symptoms of patients and a skin prick test were used. A skin patch test was used to determine corticosteroid hypersensitivity. Total IgE values and total eosinophil count were obtained for all patients. Total symptom scores were calculated for the severity of symptoms and to determine the response to therapy using intranasal corticosteroids. Results: the incidence of corticosteroid hypersensitivity determined via the skin patch test was 14.0% (21 out of 150 patients). A difference was observed for patch test positivity results between the study and control groups (14% vs. 0%, respectively). Serum IgE levels and total eosinophil count were higher among patients who had corticosteroid hypersensitivity (p:0.005 and p:0.004, respectively). Patients unresponsive to intranasal corticosteroids had a higher incidence of corticosteroid hypersensitivity (71.4% vs. 4.4%, p<0.001). Conclusion: our study is the largest to date investigating CH in patients with allergic rhinitis and patients with allergic rhinitis have been found to have a high incidence (14%) of corticosteroid hypersensitivity, which may affect the response of patients to intranasal corticosteroid treatment

Theoretical Limits on Time Delay Estimation for Ultra-Wideband Cognitive Radios

In this paper, theoretical limits on time delay estimation are studied for ultra-wideband (UWB) cognitive radio systems. For a generic UWB spectrum with dispersed bands, the Cramer-Rao lower bound (CRLB) is derived for unknown channel coefficients and carrier-frequency offsets (CFOs). Then, the effects of unknown channel coefficients and CFOs are investigated for linearly and non-linearly modulated training signals by obtaining specific CRLB expressions. It is shown that for linear modulations with a constant envelope, the effects of the unknown parameters can be mitigated. Finally, numerical results, which support the theoretical analysis, are presented.Comment: IEEE ICUWB 200

Activated GTPase movement on an RNA scaffold drives co-translational protein targeting

Approximately one-third of the proteome is initially destined for the eukaryotic endoplasmic reticulum or the bacterial plasma membrane. The proper localization of these proteins is mediated by a universally conserved protein-targeting machinery, the signal recognition particle (SRP), which recognizes ribosomes carrying signal sequences and, through interactions with the SRP receptor delivers them to the protein-translocation machinery on the target membrane. The SRP is an ancient ribonucleoprotein particle containing an essential, elongated SRP RNA for which precise functions have remained elusive. Here we used single-molecule fluorescence microscopy to show that the Escherichia coli SRP–SRP receptor GTPase complex, after initial assembly at the tetraloop end of SRP RNA, travels over 100 Å to the distal end of this RNA, where rapid GTP hydrolysis occurs. This movement is negatively regulated by the translating ribosome and, at a later stage, positively regulated by the SecYEG translocon, providing an attractive mechanism for ensuring the productive exchange of the targeting and translocation machineries at the ribosome exit site with high spatial and temporal accuracy. Our results show that large RNAs can act as molecular scaffolds that enable the easy exchange of distinct factors and precise timing of molecular events in a complex cellular process; this concept may be extended to similar phenomena in other ribonucleoprotein complexes

Prognosis of a case with paresthesia associated with prolonged touching of an endodontic paste to the inferior alveolar nerve

Paresthesia is described as an abnormal sensation, such as burning, pricking, tickling, tingling, formication or numbness. Several conditions can cause paresthesia. This article presents a case of paresthesia caused by the extrusion of endodontic paste (Endomethasone®) into the mandibular canal. The clinical manifestations comprised the numbness on the right side of the mandible and right lower lip, appearing after endodontic treatment. After a mandibular block and infiltration anesthesia a mucoperiostal flap was raised and the extruded Endomethasone® was removed successfully. A therapy with antibiotic, B vitamin complex and an analgesic were prescribed. The patient reported an improvement in pain and headache after one week later and in burning after two weeks. After a four months follow-up, she became symptom free. Also sixteen months later she had any symptoms. Normalization of sensation shows that the neurotoxic effects of Endomethasone® are reversible after more than one month from the first touch of Endomethasone® to the inferior alveolar nerv