Structural Ordering of Disordered Ligand-Binding Loops of Biotin Protein Ligase into Active Conformations as a Consequence of Dehydration

Mycobacterium tuberculosis (Mtb), a dreaded pathogen, has a unique cell envelope composed of high fatty acid content that plays a crucial role in its pathogenesis. Acetyl Coenzyme A Carboxylase (ACC), an important enzyme that catalyzes the first reaction of fatty acid biosynthesis, is biotinylated by biotin acetyl-CoA carboxylase ligase (BirA). The ligand-binding loops in all known apo BirAs to date are disordered and attain an ordered structure only after undergoing a conformational change upon ligand-binding. Here, we report that dehydration of Mtb-BirA crystals traps both the apo and active conformations in its asymmetric unit, and for the first time provides structural evidence of such transformation. Recombinant Mtb-BirA was crystallized at room temperature, and diffraction data was collected at 295 K as well as at 120 K. Transfer of crystals to paraffin and paratone-N oil (cryoprotectants) prior to flash-freezing induced lattice shrinkage and enhancement in the resolution of the X-ray diffraction data. Intriguingly, the crystal lattice rearrangement due to shrinkage in the dehydrated Mtb-BirA crystals ensued structural order of otherwise flexible ligand-binding loops L4 and L8 in apo BirA. In addition, crystal dehydration resulted in a shift of ∼3.5 Å in the flexible loop L6, a proline-rich loop unique to Mtb complex as well as around the L11 region. The shift in loop L11 in the C-terminal domain on dehydration emulates the action responsible for the complex formation with its protein ligand biotin carboxyl carrier protein (BCCP) domain of ACCA3. This is contrary to the involvement of loop L14 observed in Pyrococcus horikoshii BirA-BCCP complex. Another interesting feature that emerges from this dehydrated structure is that the two subunits A and B, though related by a noncrystallographic twofold symmetry, assemble into an asymmetric dimer representing the ligand-bound and ligand-free states of the protein, respectively. In-depth analyses of the sequence and the structure also provide answers to the reported lower affinities of Mtb-BirA toward ATP and biotin substrates. This dehydrated crystal structure not only provides key leads to the understanding of the structure/function relationships in the protein in the absence of any ligand-bound structure, but also demonstrates the merit of dehydration of crystals as an inimitable technique to have a glance at proteins in action

The Physiological Roles of Rhopr-kinins and the Molecular Characterization of their Gene in the Blood-gorging Insect, Rhodnius prolixus

The dramatic feeding-related activities of the Chagas' disease vector, Rhodnius prolixus are under neurohormonal regulation of serotonin and various neuropeptides. One such family of neuropeptides, the insect kinins, possesses diuretic, digestive and myotropic activities in many insects. In R. prolixus, they co-localize with the corticotropin-releasing factor (CRF)-like diuretic hormone (DH) in neurosecretory cell bodies and their abdominal neurohaemal sites. Additionally, kinins are present in endocrine cells of the midgut and are known to stimulate hindgut and midgut contractions. Through the experimentation presented in this dissertation, the cloning and spatial expression of the R. prolixus kinin (Rhopr-kinin) transcript is described. Physiological bioassays demonstrate the myostimulatory effects of selected Rhopr-kinin peptides and also illustrate the augmented responses of hindgut contractions to co-application of Rhopr-kinin and Rhopr-CRF/DH. The irreversible effects of two synthetic kinin analogs on the hindgut relative to the native kinins also exhibit the prospective biotechnological significance of this study.MAS

Harnessing the potential of tobacco cessation programme amidst COVID-19 pandemic in India

The COVID-19 pandemic of the 21st Century continues to spread, and tobacco users are at a higher risk of contracting the disease. As a measure to contain its spread, many nations have called for various measures like maintaining social distancing norms, the prohibition of spitting in the public place, partial or complete lockdown, and many more. This shutdown episode has disrupted the entire supply chain in our country, and it is quite natural that tobacco users are also experiencing a scarcity of tobacco products, like others. This adverse situation is an opportune moment for the Indian health systems to target tobacco users to motivate, facilitate, and support the cessation process. Additionally, social distancing can be achieved by utilizing and optimizing our existing health services. In our country, we have dedicated regional & national quitlines and m-Cessation facilities for tobacco users who are willing to quit. These initiatives could reduce the risk of COVID among tobacco users, facilitate the tobacco cessation movement, and provide credence to the advocacy for increasing taxes on tobacco products in the country

Ralstonia solanacearum Iron Scavenging by the Siderophore Staphyloferrin B Is Controlled by PhcA, the Global Virulence Regulator

PhcA is a transcriptional regulator that activates expression of multiple virulence genes in the plant pathogen Ralstonia solanacearum. Relative to their wild-type parents, phcA mutants overproduced iron-scavenging activity detected with chrome azurol S siderophore detection medium. Transposon mutagenesis of strain AW1-PC (phcA1) generated strain GB6, which was siderophore negative but retained weak iron-scavenging activity. The ssd gene inactivated in GB6 encodes a protein similar to group IV amino acid decarboxylases, and its transcription was repressed by iron(III) and PhcA. ssd is the terminal gene in a putative operon that also appears to encode three siderophore synthetase subunits, a integral membrane exporter, and three genes with no obvious role in siderophore production. A homologous operon was found in the genomes of Ralstonia metallidurans and Staphylococcus aureus, both of which produce the polycarboxylate siderophore staphyloferrin B. Comparison of the siderophores present in culture supernatants of R. solanacearum, R. metallidurans, and Bacillus megaterium using chemical tests, a siderophore utilization bioassay, thin-layer chromatography, and mass spectroscopy indicated that R. solanacearum produces staphyloferrin B rather than schizokinen as was reported previously. Inactivation of ssd in a wild-type AW1 background resulted in a mutant almost incapable of scavenging iron but normally virulent on tomato plants. AW1 did not produce siderophore activity when cultured in tomato xylem sap, suggesting that the main location in tomato for R. solanacearum during pathogenesis is iron replete

Exposure to second hand smoke and its correlates in Northern State of India

Background: Second-hand smoke (SHS) has enormous adverse health impacts with grave health implications for the next generation. Section 4 of Indian legislation, Cigarettes and Other Tobacco Products Act, prohibits smoking at public places, thus protecting people from SHS. Objective: The objective of present study was to assess the exposure to SHS at home and working areas in Punjab, India. Methods: The present cross-sectional study was conducted from December 2015 to March 2016. A three-stage sampling technique was used for collecting data from three randomly selected districts representing three major regions of Punjab, India. A sample size of 510 individuals was divided equally into an urban and rural area with proportionate sampling on the basis of subsets of age groups and gender. The questionnaire based on tobacco questions for the survey, a subset of key questions from global adult tobacco survey was used. Results: At home, the odds of exposure to SHS exposure was higher (odds ratio [OR] = 2.2, 95% confidence interval [CI] = 0.6–7.3) in urban area, females, low educational status, and nongovernment employee as compared to their counterparts. At workplace, (OR = 3.9 and 95% CI = 1.11–14.3) SHS exposure was higher in rural area, among males, primary and middle education and nongovernment or self-employed occupation. Conclusion: SHS exposure was low in Punjab, India especially in females as compared to other states of the country. The socio-economic disadvantaged groups and people with low education were more likely to experience exposure to SHS at workplace, which should be targeted to reduce tobacco consumption

Awareness of electronic cigarettes in India: Findings from the 2016–2017 Global Adult Tobacco Survey (GATS)

Electronic nicotine delivery systems (ENDS) have recently emerged as a public health threat globally. Despite the low proportion of e-cigarette users (1.22%) reported in the Global Adult Tobacco Survey-2, the Government of India enacted the Prohibition of E-cigarettes Act 2019 (PECA), prohibiting all forms of ENDS/ENNDS. The current analysis presents nationally representative findings on the level of awareness of e-cigarettes in India and its correlates and characteristics of those aware of e-cigarettes. Methods: The current secondary analysis from GATS-2 among adults aged 15 years and above from all states and Union Territories (UTs) of India used a standard protocol for data collection & management. A multi-stage cluster sampling design was used. The respondents who were aware of e-cigarettes were included (n=2524). Binomial Logistic regression analysis was conducted, and adjusted odds ratios (aOR), with 95% CI, were calculated to measure the associations between independent and dependent variables. Results: 3.4% of the respondents were aware (either heard or seen) of e-cigarettes and their awareness was found significantly higher among males (aOR=2.07; 95%CI= 1.90-2.24), urban population (aOR=2.83, 95%CI=2.61-3.07), higher education (aOR=0.41, 95%CI=0.38-0.45). Conclusion: Public awareness campaigns about the harms of e-cigarettes and the law (PECA) need to be rolled out in urban and rural areas. Capacity-building exercises of implementers and enforcers at the grassroots level could also support communicating the harms to hard-to-reach groups. Further, regular compliance monitoring of the legislation and prosecution of violators would facilitate its effective implementation at the national and sub-national levels. </p

Prioritizing tobacco control & its cessation under sustainable development goals with a focus on India

Sustainable development goals (SDGs) were meant to put each and everywhere 'at par'. The tobacco epidemic globally is one major deterrent to their achievement. While it gets addressed under SDG 3 through the Framework Convention on Tobacco Control (FCTC) - the World Health Organization (WHO) global treaty (the target 3.a of SDG 3), the progress made globally and by India is slow. As a result, many countries may fall short of achieving the target of reducing tobacco usage (taking 2016 as base year) by 30 per cent by the year 2030. India with its high burden of tobacco use and abysmally low quitting along with soaring economic costs of tobacco related diseases and deaths can do better with the engagement of multisectoral stakeholders to strengthen tobacco control under SDGs. Moreover, there is a need to emphasize that the goal of O - Offer to Quit of WHO MPOWER can be achieved through increasing 'onus' on policy makers, and strategists, and opportunities for masses, tobacco users, healthcare professionals (HCPs) and enforcers to have tobacco cessation delivered optimally. By doing so, the United Nations can significantly facilitate a reduction in tobacco use and the resultant economic costs. Furthermore, it will assist the WHO to fulfil the targets set for 2030 under SDG 3.a by the FCTC member countries. In addition, it will fulfil the vision and mission defined in the Chandigarh declaration of the 5th National Conference on Tobacco or Health for India to be tobacco free by 2030

Prevalence of depression and anxiety in patients with diabetes mellitus at a secondary level government health-care facility in Northern India

Background: Diabetes is one of the largest global health emergencies of the 21st century. As per the International Diabetes Federation (IDF) in 2020, 463 million people have diabetes in the world and 88 million people in the Southeast Asia region. Seventy-seven million out of 88 million belong to India. The prevalence of diabetes in the population is 8.9%, according to the IDF. There exists a bidirectional association between diabetes and depression/anxiety, with both worsening each other's prognosis. The prevalence rates of depression could be up to three times higher in patients with type 1 diabetes and twice as high in people with type 2 diabetes, while anxiety disorders are seen in 40% of the patients with type 1 or 2 diabetes mellitus. Although no single cause has been identified for the positive correlation between diabetes and depression and anxiety, it is widely accepted that hypothalamic–pituitary–adrenal dysfunction, pro-inflammatory cytokines, and a host of other epigenetic factors are responsible for the increased prevalence. This study was undertaken to check the prevalence of anxiety and depression in patients with diabetes mellitus attending the outdoor clinics of internal medicine department at a district hospital, Fatehgarh Sahib, Punjab. Materials and Methods: Four hundred patients were included in our study. A semi-structured pro forma was used to collect demographic details, and anxiety and depression were screened using the Hamilton Anxiety and Depression Rating Scales, respectively. Data collected were subjected to statistical analysis. Results: Fifty-eight percent of the sample population had depression and 27% had anxiety, with a vast majority having mild symptoms. Gender and duration of diabetes were statistically significant variables affecting prevalence and severity, with females having a higher mean scores of depression and anxiety than males and higher scores with greater duration of the illness. Conclusions: This study implies need for routine screening of depression and anxiety symptoms in patients with diabetes mellitus and treating them at the earliest