Prokaryotic DNA Crossroads: Holliday Junction Formation and Resolution.

Cells are continually exposed to a multitude of internal and external stressors, which give rise to various types of DNA damage. To protect the integrity of their genetic material, cells are equipped with a repertoire of repair proteins that engage in various repair mechanisms, facilitated by intricate networks of protein-protein and protein-DNA interactions. Among these networks is the homologous recombination (HR) system, a molecular repair mechanism conserved in all three domains of life. On one hand, HR ensures high-fidelity, template-dependent DNA repair, while on the other hand, it results in the generation of combinatorial genetic variations through allelic exchange. Despite substantial progress in understanding this pathway in bacteria, yeast, and humans, several critical questions remain unanswered, including the molecular processes leading to the exchange of DNA segments, the coordination of protein binding, conformational switching during branch migration, and the resolution of Holliday Junctions (HJs). This Review delves into our current understanding of the HR pathway in bacteria, shedding light on the roles played by various proteins or their complexes at different stages of HR. In the first part of this Review, we provide a brief overview of the end resection processes and the strand-exchange reaction, offering a concise depiction of the mechanisms that culminate in the formation of HJs. In the latter half, we expound upon the alternative methods of branch migration and HJ resolution more comprehensively and holistically, considering the historical research timelines. Finally, when we consolidate our knowledge about HR within the broader context of genome replication and the emergence of resistant species, it becomes evident that the HR pathway is indispensable for the survival of bacteria in diverse ecological niches.

Global genome and transcription-coupled nucleotide excision repair pathway in prokaryotes.

In all cells-from bacteria to humans-nucleotide excision repair (NER) is a highly conserved, versatile DNA repair pathway that is responsible for the removal of a wide variety of DNA helix-distorting lesions arising from both endogenous and exogenous sources. In many organisms including bacteria, fungi, animals, and plants, NER occurs through two sub-pathways: the global genome NER (GG-NER) pathway and the transcription- coupled NER (TC-NER) pathway. Although essential factors and basic steps involved in NER have been identified, mechanisms and stages of their assembly process are not well understood. In this review, we summarize recent literature about protein interaction networks that manifest during initial stages of bacterial NER pathway while highlighting some of the key functional studies.

Efficacy and Safety of the Anti-Mucosal Addressin Cell Adhesion Molecule-1 Antibody Ontamalimab in Patients with Moderate-to-Severe Ulcerative Colitis or Crohn's Disease.

BACKGROUND AND AIMS: Ontamalimab is a fully human immunoglobulin G2 monoclonal antibody against mucosal addressin cell adhesion molecule-1 developed as treatment for inflammatory bowel disease. METHODS: Six phase 3, multicentre, randomised, double-blind, placebo-controlled clinical trials compared efficacy and safety of ontamalimab [25 mg and 75 mg once every 4 weeks] with placebo in patients with moderate-to-severe ulcerative colitis or Crohn's disease [two induction studies and one re-randomised maintenance study per condition]. This clinical trial programme was discontinued in 2020 for reasons unrelated to drug safety/efficacy; Crohn's disease studies are described in the supplementary materials. RESULTS: The induction [12-week] and maintenance [52-week] studies included 659 and 366 randomised patients, respectively. More patients who received ontamalimab induction than placebo achieved the primary endpoint of clinical remission at week 12 [25 mg, 18.5% vs 15.8% (p = 0.617), 27.0% vs 12.5% (p = 0.027); 75 mg, 29.8% vs 15.8% (p = 0.018), 29.5% vs 12.5% (p = 0.014)]; significantly more patients who received ontamalimab maintenance therapy than placebo achieved week 52 clinical remission [25 mg, 53.5% vs 8.2%, p < 0.001; 75 mg, 40.2% vs 12.8%, p < 0.001]. Endoscopic improvement was generally significantly different vs placebo [induction: 25 mg, 27.8% vs 21.1 (p = 0.253), 35.1% vs 12.5% (p = 0.001); 75 mg, 41.1% vs 21.1 (p = 0.002), 33.9% vs 12.5% (p = 0.003); maintenance: 25 mg, 56.3% vs 9.6% (p < 0.001); 75 mg, 48.8% vs 15.1% (p < 0.001)]. Adverse event rates were similar between ontamalimab and placebo groups. CONCLUSIONS: Ontamalimab 75 mg was effective with no safety concerns as induction and maintenance therapy for patients with moderate-to-severe ulcerative colitis.

Safety and efficacy of multimatrix mesalamine in paediatric patients with mild-to-moderate ulcerative colitis: a phase 3, randomised, double-blind study.

Background: Previous studies have demonstrated the tolerability and efficacy of multimatrix mesalamine in inducing and maintaining remission in adults with mild-to-moderate ulcerative colitis (UC). We evaluated the safety and efficacy of low-dose and high-dose once-daily multimatrix mesalamine in children and adolescents with mild-to-moderate UC or those in remission. Methods: This prospective, randomised, parallel-group, phase 3 study (8-week double-blind acute [DBA] phase; 26-week double-blind maintenance [DBM] phase; and an additional 8-week, open-label acute [OLA] phase) was conducted in 33 sites across North America, Europe, and the Middle East between December 12, 2014, and November 28, 2018. Eligible patients aged 5-17 years and weighing 18-90 kg were randomised 1:1 to either low (900-2400 mg) or high (1800-4800 mg) oral doses of multimatrix mesalamine once daily, stratified by body weight. Interactive response technology was used for randomisation. The primary efficacy outcome was to estimate the clinical response of multimatrix mesalamine (two doses) in different weight groups. Efficacy and safety analyses were conducted in the safety analysis set (Clinicaltrials.gov: NCT02093663; Study completed). Findings: Overall, 107 patients were randomised into the DBA (n = 54) or DBM phase (n = 88; directly or after completing the double-blind or OLA phases); the overall safety analysis set included 105 patients. In the DBA phase, the high-dose group (n = 17; 65.4%) achieved a higher clinical response rate than the low-dose (n = 10; 37.0%) group; difference 28.3% (95% CI: 2.5-54.2; p = 0.039), odds ratio (OR) 3.21 (95% CI: 1.04-9.88). In the DBM phase at Week 26, similar proportions of patients maintained clinical response in the low-dose (n = 23; 54.8%) and high-dose (n = 24; 53.3%) groups: OR 0.99 (0.42-2.34); p = 0.981. Overall, 246 treatment-emergent adverse events (TEAEs) were reported in 73 patients (69.5%); 23 TEAEs in 14 patients (13.3%) were considered related to the study drug. No treatment-related deaths were reported. Interpretation: Our findings suggested that the benefit-risk ratio of once-daily multimatrix mesalamine in paediatric patients was favourable and comparable with that reported in adults with mild-to-moderate UC. Funding: Shire Development LLC, a Takeda company.

SUMOylation of yeast Pso2 enhances its translocation and accumulation in the mitochondria and suppresses methyl methanesulfonate-induced mitochondrial DNA damage.

Saccharomyces cerevisiae Pso2/SNM1 is essential for DNA interstrand crosslink (ICL) repair; however, its mechanism of action remains incompletely understood. While recent work has revealed that Pso2/Snm1 is dual-localized in the nucleus and mitochondria, it remains unclear whether cell-intrinsic and -extrinsic factors regulate its subcellular localization and function. Herein, we show that Pso2 undergoes ubiquitination and phosphorylation, but not SUMOylation, in unstressed cells. Unexpectedly, we found that methyl methanesulfonate (MMS), rather than ICL-forming agents, induced robust SUMOylation of Pso2 on two conserved residues, K97 and K575, and that SUMOylation markedly increased its abundance in the mitochondria. Reciprocally, SUMOylation had no discernible impact on Pso2 translocation to the nucleus, despite the presence of steady-state levels of SUMOylated Pso2 across the cell cycle. Furthermore, substitution of the invariant residues K97 and K575 by arginine in the Pso2 SUMO consensus motifs severely impaired SUMOylation and abolished its translocation to the mitochondria of MMS-treated wild type cells, but not in unstressed cells. We demonstrate that whilst Siz1 and Siz2 SUMO E3 ligases catalyze Pso2 SUMOylation, the former plays a dominant role. Notably, we found that the phenotypic characteristics of the SUMOylation-defective mutant Pso2K97R/K575R closely mirrored those observed in the Pso2Δ petite mutant. Additionally, leveraging next-generation sequencing analysis, we demonstrate that Pso2 mitigates MMS-induced damage to mitochondrial DNA (mtDNA). Viewed together, our work offers previously unknown insights into the link between genotoxic stress-induced SUMOylation of Pso2 and its preferential targeting to the mitochondria, as well as its role in attenuating MMS-induced mtDNA damage.

Design of a phase 3, randomized, double-blind, placebo-controlled study to evaluate the efficacy and safety of prucalopride in pediatric patients with functional constipation.

Background: A previous phase 3 trial of prucalopride in pediatric patients (6 months-18 years old) with functional constipation (FC) demonstrated no efficacy versus placebo. We designed an additional phase 3 trial to further assess the efficacy, long-term safety and tolerability of prucalopride in children and adolescents. Methods: This multicenter trial (ClinicalTrials.gov identifier: NCT04759833; EudraCT number: 2022-003221-22) comprises a 12-week, randomized, double-blind, placebo-controlled phase, followed by a 36-week, double-blind, safety extension phase. Approximately 240 toilet-trained patients aged 3-17 years will be randomized 1:1:1 to receive low- (0.04 mg/kg) or high-dose (0.08 mg/kg) prucalopride, or placebo once daily. Fifteen non-toilet-trained patients ≥6 months old with FC will be included in an exploratory efficacy and safety analysis. Discussion: The efficacy endpoints used in this study will differ from those used in adults and in the previous pediatric phase 3 trial; they have been adapted to be more suitable for a wider age range of pediatric patients. Both study phases will be longer than in the previous pediatric study, providing a longer time period in which to assess the efficacy and safety of prucalopride. Study participants will be identified using the modified Rome IV criteria for FC, instead of the Rome III criteria, and non-toilet-trained patients will be included, which will broaden the population of pediatric patients assessed. Patients will undergo fecal disimpaction before randomization and undergo standardized continuous behavioral therapy throughout the trial. This pediatric study of prucalopride will aim to demonstrate the efficacy and long-term safety of this treatment.

Modified truncated Hochberg procedure for multiple endpoints: An application in a confirmatory trial for pediatric functional constipation.

BACKGROUND: In confirmatory clinical trials, it is critical to have appropriate control of multiplicity for multiple comparisons or endpoints. When multiplicity-related issues arise from different sources (e.g., multiple endpoints, multiple treatment arms, multiple interim data-cuts and other factors), it can become complicated to control the family-wise type I error rate (FWER). Therefore, it is crucial for statisticians to fully understand the multiplicity adjustment methods and the objectives of the analysis regarding study power, sample size and feasibility in order to identify the proper multiplicity adjustment strategy. METHODS: In the context of multiplicity adjustment of multiple dose levels and multiple endpoints in a confirmatory trial, we proposed a modified truncated Hochberg procedure in combination with a fixed-sequence hierarchical testing procedure to strongly control the FWER. In this paper, we provided a brief review of the mathematical framework of the regular Hochberg procedure, the truncated Hochberg procedure and the proposed modified truncated Hochberg procedure. An ongoing phase 3 confirmatory trial for pediatric functional constipation was used as a real case application to illustrate how the proposed modified truncated Hochberg procedure will be implemented. A simulation study was conducted to demonstrate that the study was adequately powered and the FWER was strongly controlled. CONCLUSION: This work is expected to facilitate the understanding and selection of adjustment methods for statisticians.

Macrophage activation highlight an important role for NER proteins in the survival, latency and multiplication of Mycobacterium tuberculosis.

Nucleotide excision repair (NER) is one of the most extensively studied DNA repair processes in both prokaryotes and eukaryotes. The NER pathway is a highly conserved, ATP-dependent multi-step process involving several proteins/enzymes that function in a concerted manner to recognize and excise a wide spectrum of helix-distorting DNA lesions and bulky adducts by nuclease cleavage on either side of the damaged bases. As such, the NER pathway of Mycobacterium tuberculosis (Mtb) is essential for its survival within the hostile environment of macrophages and disease progression. This review focuses on present published knowledge about the crucial roles of Mtb NER proteins in the survival and multiplication of the pathogen within the macrophages and as potential targets for drug discovery.

Interrogating the substrate specificity landscape of UvrC reveals novel insights into its non-canonical function.

Although it is relatively unexplored, accumulating data highlight the importance of tripartite crosstalk between nucleotide excision repair (NER), DNA replication, and recombination in the maintenance of genome stability; however, elucidating the underlying mechanisms remains challenging. While Escherichia coli uvrA and uvrB can fully complement polAΔ cells in DNA replication, uvrC attenuates this alternative DNA replication pathway, but the exact mechanism by which uvrC suppresses DNA replication is unknown. Furthermore, the identity of bona fide canonical and non-canonical substrates for UvrCs are undefined. Here, we reveal that Mycobacterium tuberculosis UvrC (MtUvrC) strongly binds to, and robustly cleaves, key intermediates of DNA replication/recombination as compared with the model NER substrates. Notably, inactivation of MtUvrC ATPase activity significantly attenuated its endonuclease activity, thus suggesting a causal link between these two functions. We built an in silico model of the interaction of MtUvrC with the Holliday junction (HJ), using a combination of homology modeling, molecular docking, and molecular dynamic simulations. The model predicted residues that were potentially involved in HJ binding. Six of these residues were mutated either singly or in pairs, and the resulting MtUvrC variants were purified and characterized. Among them, residues Glu595 and Arg597 in the helix-hairpin-helix motif were found to be crucial for the interaction between MtUvrC and HJ; consequently, mutations in these residues, or inhibition of ATP hydrolysis, strongly abrogated its DNA-binding and endonuclease activities. Viewed together, these findings expand the substrate specificity landscape of UvrCs and provide crucial mechanistic insights into the interplay between NER and DNA replication/recombination.

Seropositivity among Blood Samples Drawn from Suspected Dengue Cases at a Tertiary Care Centre of Nepal: A Descriptive Cross-sectional Study.

INTRODUCTION: The cases of dengue fever have been reported more frequently in Nepal these days. The aim of this study was to find the prevalence of seropositivity among blood samples drawn from suspected dengue cases at a tertiary care centre of Nepal. METHODS: A descriptive cross-sectional study was conducted at a tertiary care hospital from 1st June 2017 to 31st October 2018 after getting approval from the Institutional Review Committee (Reference number: 23/2016). A total of 537 suspected dengue patients were selected for the study using convenience sampling. These dengue positive sera were assayed for their reactivity with Immunoglobulin M and Immunoglobulin G present in sera and synthetic peptides of dengue virus by enzyme linked immunosorbent assay. Data was entered and analysed in Microsoft Excel 2016. Point estimate at 95% Confidence Interval was calculated along with frequency, percentage, mean and standard deviation. RESULTS: Among 537 suspected dengue cases, the seropositivity for dengue was found in 124 (23.09%) (19.52-26.65 at 95% Confidence Interval) of the serum. CONCLUSIONS: The present study revealed that dengue was more prevalent in our setting compared to similar studies. All the synthetic peptides showed reactivity with dengue-positive sera with maximum reactivity shown by RR2 peptide. In dengue-positive sera, RR2 peptide of dengue virus identified more Immunoglobulin M than Immunoglobulin G.

Dynamic contrast-enhanced magnetic resonance perfusion volumetrics can differentiate tuberculosis of the spine and vertebral malignancy.

BACKGROUND: There is considerable overlap in radiologic features of tubercular and malignant spinal lesions on conventional magnetic resonance imaging (MRI). PURPOSE: To evaluate the role of dynamic contrast-enhanced (DCE) MRI perfusion parameters in differentiating vertebral malignancy from spinal tuberculosis. MATERIAL AND METHODS: This was a prospective study and we enrolled consecutive patients presenting with a clinical/radiologic evidence of vertebral lesions. DCE-MRI of the spine was performed using 3D volume interpolated breath-hold examination (VIBE) sequence after intravenously injecting 0.1 mmol/kg body weight of gadopentetate dimeglumine. We used Tofts model to calculate DCE parameters that included Ktrans (transfer constant), kep (rate constant), ve (fractional volume of extracellular extravascular space), and iAUC (initial area under the curve). We compared the mean value of each perfusion parameter by type of lesion (tubercular/malignant) at 0.05 significance level and performed receiver operating characteristic curve analysis. RESULTS: We could confirm histologic/cytologic diagnosis in 35 of the 45 patients recruited. Of these, 19 were tubercular and 16 were malignant lesions. The mean (± standard deviation) of kep (min-1) was significantly higher (2.89 ± 3.3) in malignant compared to tubercular lesions (0.81 ± 0.19), whereas ve was significantly lower in malignant (0.27 ± 0.13 mL/g) compared to benign lesions (0.47 ± 0.12 mL/g) at 0.05 significance level. kep cutoff of ≥1.17 min-1 had a sensitivity of 93.8% and specificity of 100% with a diagnostic accuracy of 94.4% in detecting malignant disease. CONCLUSION: High kep is the single best predictor of malignant vertebral lesions. We recommend kep cutoff value of ≥1.17 min-1 that has high diagnostic accuracy in identifying malignant lesions.

Influence of Implant Angulation and Implant Number on the Accuracy of Definitive Casts.

BACKGROUND: The present study determined the effect of implant angulation and implant number on the dimensional precision of implant definitive casts. MATERIALS AND METHODS: Three definitive casts with implant analogs placed in a triangular pattern were made from dental stone. Group I was control group, Group II in which implant numbers 1 and 3 were at 5° convergence to the implant number 2. Group III in which implant numbers 1 and 3 are at 5° divergence to the implant number 2. Group IV in which implant numbers 1 and 3 are at 10° convergence to the implant number 2. Implant analogs were secured in all the definitive casts with cyanoacrylate. Three open tray impressions of the definitive cast were obtained and poured in Type IV dental stone. Coordinates in the three planes were measured at implant analog top surface and base of the cast using a fine tip measuring stylus. The data were aligned and angular differences between implant analog vectors from definitive and duplicate casts were measured. RESULTS: There was a significant correlation between dental implant number and dental implant angulation (P < 0.05). There was a significant result when comparing the effect of the parallel group from 5° divergence and 10° convergence groups. A significant results while comparing the parallel group with 5° divergence and 10° convergence was obtained whereas while comparing 5° divergence, 5° divergence, and 10° convergence a nonsignificant difference was obtained (P > 0.05). CONCLUSION: Close proximity of implant angulation toward right-angled direction results in higher precision of implant.

Assessment of Titanium Level in Submucosal Plaque Around Healthy Implants and Implants with Peri-implantitis: A Clinical Study.

BACKGROUND: The present study focused on assessing the level of titanium in submucosal plaque in the peri-implant area with peri-implantitis in comparison to healthy implants. METHODOLOGY: Sixty patients with titanium dental implants were recruited. The degree of titanium in submucosal plaque around peri-implantitis and healthy implants was estimated using inductively coupled plasma mass spectrometry. RESULTS: The mean ± standard deviation probing depth in Group I was 3.12 ± 1.1 and in Group II was 7.2 ± 2.5; gingival index was 0.64 ± 0.3 and 1.64 ± 0.8 in Group I and Group II, respectively. The plaque index was 0.82 ± 0.2 in Group I and 1.5 ± 0.6 in Group II. The mean plaque mass in Group I was 24.1 ± 3.8 ng/ul and 49.3 ± 6.4 ng/ul in Group II. The mean titanium level in Group I was 0.08 ± 0.02 µg and in Group II was 0.91 ± 0.04 µg. A highly significant difference between both groups was found (P < 0.05). CONCLUSION: There was a significantly higher titanium level in submucosal plaque around dental implants with signs of peri-implantitis as compared to healthy dental implants.

Evaluation of Quality of Endodontic Re-Treatment and Changes in Periapical Status.

BACKGROUND: The present study was conducted to assess quality of root canal (RC) filling before and after RC re-treatment. MATERIALS AND METHODS: Two hundred and thirty-eight radiographs of failed endodontic treatment were assessed. The periapical status of the endodontic treatment was evaluated with periapical index (PAI) scoring system. PAI <3 showed absence and PAI >3 showed presence of periapical lesion. RESULTS: There was a statistically significant increase in scores 1 and 3 and decrease in scores 2, 4, 5, and 6 after treatment (P < 0.05). PAI score >3 was seen in 37% before which decreased to 16% after endodontic retreatment. 34.6% obturation was homogenous and 65.4% was nonhomogenous before endodontic retreatment. After endodontic retreatment, 95.2% became homogenous and 4.8% nonhomogenous. The reason for endodontic failure was furcation in 2%, iatrogenic causes in 3%, loss of coronal seal in 16%, periapical pathology in 25%, and inadequate root filling in 54%. CONCLUSION: There was significant improvement and decrease in size of periapical lesions in re-endodontic cases as compared to primary RC treated teeth.

Spontaneous subcapsular renal haematoma in a diabetic patient with acute pyelonephritis.

Spontaneous renal subcapsular haematoma is not a common clinical condition. It is mainly caused by renal tumours, vascular diseases, trauma, anticoagulation, and urological interventions. Patients present with sudden onset abdominal pain mainly in the flank region, haematuria, and abdominal tenderness. We report a case of 42-year-old diabetic patient who presented with a history of fever and left flank pain with clinical signs of pyelonephritis, on whom abdominal ultrasound and computed tomography demonstrated a subcapsular renal haematoma which was confirmed by percutaneous aspiration. Management with antibiotics and percutaneous nephrostomy proved successful.

Novel insights into ATP-Stimulated Cleavage of branched DNA and RNA Substrates through Structure-Guided Studies of the Holliday Junction Resolvase RuvX.

Much of our understanding of the homologous recombination (HR) machinery hinges on studies using Escherichia coli as a model organism. Interestingly enough, studies on the HR machinery in different bacterial species casts doubt on the universality of the E. coli paradigm. The human pathogen Mycobacterium tuberculosis encodes two Holliday junction (HJ)-resolvase paralogues, namely RuvC and RuvX; however, insights into their structural features and functional relevance is still limited. Here, we report on structure-guided functional studies of the M. tuberculosis RuvX HJ resolvase (MtRuvX). The crystalline MtRuvX is a dimer in the asymmetric unit, and each monomer has a RNAse H fold vis-à-vis RuvC-like nucleases. Interestingly, MtRuvX also contains some unique features, including the residues essential for ATP binding/coordination of Mg2+ ions. Indeed, MtRuvX exhibited an intrinsic, robust ATPase activity, which was further accentuated by DNA cofactors. Structure-guided substitutions of single residues at the ATP binding/Mg2+coordination sites while markedly attenuating the ATPase activity completely abrogated HJ cleavage, indicating an unanticipated relationship between ATP hydrolysis and DNA cleavage. However, the affinity of ATPase-deficient mutants for the HJ was not impaired. Contrary to RuvC, MtRuvX exhibits relaxed substrate specificity, cleaving a variety of branched DNA/RNA substrates. Notably, ATP hydrolysis plays a regulatory role, rendering MtRuvX from a canonical HJ resolvase to a DNA/RNA non-sequence specific endonuclease, indicating a link between HJ resolvase and nucleic acid metabolism. These findings provide novel insights into the structure and dual-functional activities of MtRuvX, and suggest that it may play an important role in DNA/RNA metabolism.

To Evaluate the Marginal Adaptation of Porcelain Fused to Metal Crown with Different Base Metal Alloys.

INTRODUCTION: The success of any restoration depends on the marginal seal. The adaptation of castings, luting cement, and the surface structures of the margins are all important factors in achieving marginal seal. AIM: The aim of this study was to evaluate the vertical marginal discrepancy of cast copings obtained by employing conventional casting technique with two different base metal alloys with two different finish lines before and after porcelain firing. MATERIAL AND METHODS: A total of forty wax copings were fabricated with stainless steel die assembly and divided into four groups with ten specimens for each metal and each finish line. Measurements were recorded from coping margin to the stainless steel die margin for vertical marginal gap recordings. Each metal coping was finished, and porcelain application was completed. Copings with porcelain were placed on their respective dies, again subjected to the same measuring microscope for checking the vertical marginal discrepancy by the same operator and results. RESULTS: The results of the present study showed that the mean vertical marginal gaps of all the cast copings obtained in each group (G1-G8) were within clinically acceptable limits. The mean vertical marginal gap of G1 was 135.36 ± 2.30 µm, G2 was 67.22 ± 4.25 µm, G3: 39.47 ± 2.98 µm, G4: 71.00 ± 3.97 µm, G5: 109.57 ± 2.98 µm, G6: 109.57 ± 2.98 µm, and G8: 114.58 ± 2.40 µm. CONCLUSION: The difference in the vertical marginal gap of cast copings obtained in different groups was statistically highly significant at 0.005 level, while the difference in the vertical marginal gap of cast copings obtained at different points was statistically nonsignificant.

To Evaluate the Prevalence of Edentulousness, the Present Prosthetic Status, and the Need for Prosthetic Treatment in the Population of Garhwa, Jharkhand.

AIM: This study aimed to evaluate the prevalence of edentulousness, the present prosthetic status, and the need for prosthetic treatment in the population of Garhwa, Jharkhand. MATERIALS AND METHODS: A descriptive cross-sectional study was conducted among the population of Garhwa, Jharkhand. The sample size included 460 subjects belonging to an age group of 25 years and above. Detailed case history and thorough dental examination was done and all the findings were recorded and compiled. Each subject was interrogated with the help of a predesigned closed ended questionnaire. RESULTS: A total of six patients were completely edentulous, while 55.04% of males and 42.80% of females were partially edentulous. All patients who were completely edentulous were denture wearer, while among partially edentulous patients, 6.31% of males and 3.15% of females had RPD and 13.68% and 6.31% of males and females, respectively, had FPD as prosthesis. Prosthetic status of females was better than males In a comparison of prosthetic status between male and female, out of 460 subjects, 195 subjects opted for prosthesis, in which 117 were male and 78 were female. CONCLUSION: Community-based oral health education programs should be conducted to improve patients' oral health knowledge. Educating patients regarding the limitations of prosthesis as mechanical substitutes for natural teeth must be a continuing process from the initial patient contact until adjustments are completed.

The intrinsic ATPase activity of Mycobacterium tuberculosis UvrC is crucial for its damage-specific DNA incision function.

To ensure genome stability, bacteria have evolved a network of DNA repair mechanisms; among them, the UvrABC-dependent nucleotide excision repair (NER) pathway is essential for the incision of a variety of bulky adducts generated by exogenous chemicals, UV radiation and by-products of cellular metabolism. However, very little is known about the enzymatic properties of Mycobacterium tuberculosis UvrABC excinuclease complex. Furthermore, the biochemical properties of Escherichia coli UvrC (EcUvrC) are not well understood (compared to UvrA and UvrB), perhaps due to its limited availability and/or activity instability in vitro. In addition, homology modelling of M. tuberculosis UvrC (MtUvrC) revealed the presence of a putative ATP-binding pocket, although its function remains unknown. To elucidate the biochemical properties of UvrC, we constructed and purified wild-type MtUvrC and its eight variants harbouring mutations within the ATP-binding pocket. The data from DNA-binding studies suggest that MtUvrC exhibits high-affinity for duplex DNA containing a bubble or fluorescein-dT moiety, over fluorescein-adducted single-stranded DNA. Most notably, MtUvrC has an intrinsic UvrB-independent ATPase activity, which drives dual incision of the damaged DNA strand. In contrast, EcUvrC is devoid of ATPase activity; however, it retains the ability to bind ATP at levels comparable to that of MtUvrC. The ATPase-deficient variants map to residues lining the MtUvrC ATP-binding pocket. Further analysis of these variants revealed separation of function between ATPase and DNA-binding activities in MtUvrC. Altogether, these findings reveal functional diversity of the bacterial NER machinery and a paradigm for the evolution of a catalytic scaffold in UvrC.

Deciphering the essentiality and function of SxSx motif in Mycobacterium tuberculosis UvrB.

The UvrB subunit is a central component of the UvrABC incision complex and plays a pivotal role in damage recognition, strand excision and repair synthesis. A conserved structural motif (the SxSx motif) present in UvrB is analogous to a similar motif (TxGx) in the helicases of superfamily 2, whose function is not fully understood. To elucidate the significance of the SxSx (Ser143-Val144-Ser145-Cys146) motif in Mycobacterium tuberculosis UvrB (MtUvrB), different variants of MtUvrB subunit were constructed and characterized. The SxSx motif indeed was found to be essential for MtUvrB function: while Ser143 and Cys146 residues within this motif were crucial for MtUvrB function, Ser145 plays an important but less essential role. The SxSx motif-deleted mutant was drastically attenuated and three single (S143A, S145A and C146A) mutants and a double (S143A/S145A) mutant exhibited various degrees of severity in their DNA-binding, DNA helicase and ATPase activities. Taken together, these results highlight a hitherto unrecognized role for SxSx motif in the catalytic activities of UvrB.