Amino Acid Variations Research Articles

Molecular Evolution and Protein Structure Variation of Dkk Family.

Dkks have inhibitory effects on the Wnt signaling pathway, which is involved in the development of skin and its appendages and the regulation of hair growth. The nucleotide sequences were compared and analyzed to further investigate the relationship between the structure and function of the Dkk gene family and vertebrate epidermal hair. The analysis of the molecular evolution of the Dkk family revealed that the evolution rate of the genes changed significantly after speciation, with the Aves and Reptilia branches showing accelerated evolution. Additionally, positive selection was observed at specific sites. The tertiary structure of the protein was also predicted. The analysis of the functional divergence of the Dkk family revealed that the functional divergence coefficient of each gene was greater than 0, with most of the functional divergence sites were located in the Cys-2 domain and a few in the Cys-1 domain. This suggests that the amino acid and functional divergence sites may play a role in regulating the binding of the Dkk family to LRP5/6, and thus affect the inhibition of Wnt signaling, leading to different functions of Dkk1, Dkk2, and Dkk4 in the development of skin hair follicles. In addition, the Dkk families of Aves and Reptilia may have undergone adaptive evolution and functional divergence.

Isolation and Characterization of a Novel Recombinant Classical Pseudorabies Virus in the Context of the Variant Strains Pandemic in China.

Pseudorabies virus (PRV) variants were discovered in immunized pigs in Northern China and have become the dominant strains since 2011, which caused huge economic losses. In this study, a classical PRV strain was successfully isolated in a PRV gE positive swine farm. The complete genome sequence was obtained using a high-throughput sequencing method and the virus was named JS-2020. The nucleotide homology analysis and phylogenetic tree based on complete genome sequences or gC gene showed that the JS-2020 strain was relatively close to the classical Ea strain in genotype II clade. However, a large number of amino acid variations occurred in the JS-2020 strain compared with the Ea strain, including multiple immunogenic and virulence-related genes. In particular, the gE protein of JS-2020 was similar to earlier Chinese PRV strains without Aspartate insertion. However, the amino acid variations analysis based on major immunogenic and virulence-related genes showed that the JS-2020 strain was not only homologous with earlier PRV strains, but also with strains isolated in recent years. Moreover, the JS-2020 strain was identified as a recombinant between the GXGG-2016 and HLJ-2013 strains. The pathogenicity analysis proved that the PRV JS-2020 strain has typical neurogenic infections and a strong pathogenicity in mice. Together, a novel recombinant classical strain was isolated and characterized in the context of the PRV variant pandemic in China. This study provided some valuable information for the study of the evolution of PRV in China.

Mycobacterium tuberculosis β-lactamase variant reduces sensitivity to ampicillin/avibactam in a zebrafish-Mycobacterium marinum model of tuberculosis

The β-lactamase of Mycobacterium tuberculosis, BlaC, hydrolyzes β-lactam antibiotics, hindering the use of these antibiotics for the treatment of tuberculosis. Inhibitors, such as avibactam, can reversibly inhibit the enzyme, allowing for the development of combination therapies using both antibiotic and inhibitor. However, laboratory evolution studies using Escherichia coli resulted in the discovery of single amino acid variants of BlaC that reduce the sensitivity for inhibitors or show higher catalytic efficiency against antibiotics. Here, we tested these BlaC variants under more physiological conditions using the M. marinum infection model of zebrafish, which recapitulates hallmark features of tuberculosis, including the intracellular persistence of mycobacteria in macrophages and the induction of granuloma formation. To this end, the M. tuberculosis blaC gene was integrated into the chromosome of a blaC frameshift mutant of M. marinum. Subsequently, the resulting strains were used to infect zebrafish embryos in order to test the combinatorial effect of ampicillin and avibactam. The results show that embryos infected with an M. marinum strain producing BlaC show lower infection levels after treatment than untreated embryos. Additionally, BlaC K234R showed higher infection levels after treatment than those infected with bacteria producing the wild-type enzyme, demonstrating that the zebrafish host is less sensitive to the combinatorial therapy of β-lactam antibiotic and inhibitor. These findings are of interest for future development of combination therapies to treat tuberculosis.

Algae Ingestion Increases Resting and Exercised Myofibrillar Protein Synthesis Rates to a Similar Extent as Mycoprotein in Young Adults

BackgroundSpirulina [SPIR] (cyanobacterium) and chlorella [CHLO] (microalgae) are foods rich in protein and essential amino acids; however, their capacity to stimulate myofibrillar protein synthesis (MyoPS) in humans remains unknown. ObjectivesWe assessed the impact of ingesting SPIR and CHLO compared with an established high-quality nonanimal-derived dietary protein source (fungal-derived mycoprotein [MYCO]) on plasma amino acid concentrations, as well as resting and postexercise MyoPS rates in young adults. MethodsThirty-six healthy young adults (age: 22 ± 3 y; BMI: 23 ± 3 kg·m-2; male [m]/female [f], 18/18) participated in a randomized, double-blind, parallel-group trial. Participants received a primed, continuous infusion of L-[ring-2H5]-phenylalanine and completed a bout of unilateral-resistance leg exercise before ingesting a drink containing 25 g protein from MYCO (n = 12; m/f, 6/6), SPIR (n = 12; m/f, 6/6), or CHLO (n = 12; m/f, 6/6). Blood and bilateral muscle samples were collected at baseline and during a 4-h postprandial and postexercise period to assess the plasma amino acid concentrations and MyoPS rates in rested and exercised tissue. ResultsProtein ingestion increased the plasma total and essential amino acid concentrations (time effects; all P < 0.001), but most rapidly and with higher peak responses following the ingestion of SPIR compared with MYCO and CHLO (P < 0.05), and MYCO compared with CHLO (P < 0.05). Protein ingestion increased MyoPS rates (time effect; P < 0.001) in both rested (MYCO, from 0.041 ± 0.032 to 0.060 ± 0.015%·h−1; SPIR, from 0.042 ± 0.030 to 0.066 ± 0.022%·h−1; and CHLO, from 0.037 ± 0.007 to 0.055 ± 0.019%·h−1, respectively) and exercised tissue (MYCO, from 0.046 ± 0.014 to 0.092 ± 0.024%·h−1; SPIR, from 0.038 ± 0.011 to 0.086 ± 0.028%·h−1; and CHLO, from 0.048 ± 0.019 to 0.090 ± 0.024%·h−1, respectively), with no differences between groups (interaction effect; P > 0.05), but with higher rates in exercised compared with rested muscle (time × exercise effect; P < 0.001). ConclusionsThe ingestion of a single bolus of algae-derived SPIR and CHLO increases resting and postexercise MyoPS rates to a comparable extent as MYCO, despite divergent postprandial plasma amino acid responses.

The protective association of HLA-C*12:02 and HLA-DQB1*06:01 with severe acute hepatitis of unknown origin in the Japanese population.

Global surveillance has been conducted to elucidate the pathogenesis of acute hepatitis of unknown origin (AHUO), However, the factors associated with the aggravation of this serious disease are unclear. Therefore, we conducted a HLA association study to identify HLA alleles or haplotypes predisposing or protective against Japanese AHUO. The HLA 5 locus (HLA-A, HLA-B, C, DRB1, and DQB1) 4-digit genotyping results of 72 AHUO patients who underwent liver transplantation at our institution between 2000 and 2021 were compared to those of 873 healthy Japanese controls. Protective associations of HLA-B*52:01 (p-corrected (pc) = 3.15 × 10-3 ), HLA-C*12:02 (pc = 1.66 ×10-3 ), HLA-DQB1*06:01 (pc = 1.42 × 10-2 ), and HLA-DRB1*15:02 (pc = 1.36 × 10-2 ) with severe AHUO in Japanese patients were observed. The amino acid residues of tryptophan at position 156, which are located in the antigen-binding grooves of the HLA-C protein, showed a protective association with AHUO, showing a significant difference from other amino acid variations (pc = 9.0 × 10-4 ). Furthermore, 5 amino acid residues of the HLA-DQB1 protein were also protectively associated with AHUO with a significant difference from other amino acid variations (pc = 1.42 × 10-2 to 2.89 × 10-2 ). These alleles have a protective association with the aggravation of AHUO in the Japanese population.

Fluoroalkylation of Activated Allylic Acetates through Radical-Radical Coupling: Organophotoredox/DABCO Catalytic System.

A novel organophotoredox/DABCO catalytic system for the fluoroalkylation of activated allylic acetates via radical-radical coupling is described. The method offers mild reaction conditions, high selectivity, and broad substrate compatibility and enabled diverse bioactive molecules, FDA-approved drugs, and amino acid derivatives to be incorporated into transformation. This study expands the synthetic toolbox for the construction of fluorine-containing molecules.

Variability of Particulate Amino Acids and Processes Responsible for Sources into the Coastal Waters of the Bay of Bengal

Variability of Particulate Amino Acids and Processes Responsible for Sources into the Coastal Waters of the Bay of Bengal

ATP2B2 de novo variants as a cause of variable neurodevelopmental disorders that feature dystonia, ataxia, intellectual disability, behavioral symptoms, and seizures

PurposeATP2B2 encodes the variant-constrained plasma-membrane calcium-transporting ATPase-2, expressed in sensory ear cells and specialized neurons. ATP2B2/Atp2b2 variants were previously linked to isolated hearing loss in patients and neurodevelopmental deficits with ataxia in mice. We aimed to establish the association between ATP2B2 and human neurological disorders. MethodsMultinational case recruitment, scrutiny of trio-based genomics data, in silico analyses, and functional variant characterization were performed. ResultsWe assembled 7 individuals harboring rare, predicted deleterious heterozygous ATP2B2 variants. The alleles comprised 5 missense substitutions that affected evolutionarily conserved sites and 2 frameshift variants in the penultimate exon. For 6 variants, a de novo status was confirmed. Unlike described patients with hearing loss, the individuals displayed a spectrum of neurological abnormalities, ranging from ataxia with dystonic features to complex neurodevelopmental manifestations with intellectual disability, autism, and seizures. Two cases with recurrent amino-acid variation showed distinctive overlap with cerebellar atrophy-associated ataxia and epilepsy. In cell-based studies, all variants caused significant alterations in cytosolic calcium handling with both loss- and gain-of-function effects. ConclusionPresentations in our series recapitulate key phenotypic aspects of Atp2b2-mouse models and underline the importance of precise calcium regulation for neurodevelopment and cerebellar function. Our study documents a role for ATP2B2 variants in causing heterogeneous neurodevelopmental and movement-disorder syndromes.

Zwitterionic peptides: Tunable next-generation stealth nanoparticle modifications.

Adsorption of proteins to nanoparticles (NPs), a complex process that results in a protein corona, is controlled by NP surface properties that define NP interactions in vivo. Efforts to control adsorbed protein quantity through surface modification have led to improvements in circulation time or biodistribution. Still, current approaches have yet to be identified to control adsorbed protein identities within the corona. Here, we report the development and characterization of diverse zwitterionic peptides (ZIPs) for NP anti-fouling surface functionalization with specific and controllable affinity for protein adsorption profiles defined by ZIP sequence. Through serum exposure of ZIP-conjugated NPs and proteomics analysis of the resulting corona, we determined that protein adsorption profiles depend not on the exact composition of the ZIPs but on the sequence and order of charges along the sequence (charge motif). These findings pave the way for developing tunable ZIPs to orchestrate specific ZIP-NP protein adsorption profiles as a function of ZIP charge motif to better control cell and tissue specificity and pharmacokinetics and provide new tools for investigating relationships between protein corona and biological function. Furthermore, overall ZIP diversity enabled by the diversity of amino acids may ameliorate adaptive immune responses.

Genetic Characteristics and Phylogeographic Dynamics of Echovirus.

Echoviruses belong to the genus Enterovirus in the Picornaviridae family, forming a large group of Enterovirus B (EV-B) within the Enteroviruses. Previously, Echoviruses were classified based on the coding sequence of VP1. In this study, we performed a reliable phylogenetic classification of 277 sequences isolated from 1992 to 2019 based on the full-length genomes of Echovirus. In this report, phylogenetic, phylogeographic, recombination, and amino acid variability landscape analyses were performed to reveal the evolutional characteristics of Echovirus worldwide. Echoviruses were clustered into nine major clades, e.g., G1-G9. Phylogeographic analysis showed that branches G2-G9 were linked to common strains, while the branch G1 was only linked to G5. In contrast, strains E12, E14, and E16 clustered separately from their G3 and G7 clades respectively, and became a separate branch. In addition, we identified a total of 93 recombination events, where most of the events occurred within the VP1-VP4 coding regions. Analysis of amino acid variation showed high variability in the a positions of VP2, VP1, and VP3. This study updates the phylogenetic and phylogeographic information of Echovirus and indicates that extensive recombination and significant amino acid variation in the capsid proteins drove the emergence of new strains.

Comprehensive insight on protein modification by V-type agent: A chemical proteomic approach employing bioorthogonal reaction.

Organophosphorus compounds (OPs) such as chemical agents and pesticides are posing critical threats to civilians due to their irreversible phosphonylation of diverse amino acids residues forming different protein adducts. However, traditional analytical approaches are quite limited in capturing the myriad of post-translational events that affect protein functions, especially in identifying the low-abundance OP adducts. Herein a systematic proteomic strategy based on a typical click-enrich-release-identify bioorthogonal operation was firstly developed by employing an alkynyl-tagged V-type agent probe (AVP) and a biotin-based azido-enrichment linker (BTP-N3 ). AVP targeting peptides from human serum albumin (HSA) or plasma were captured by BTP-N3 via CuAAC click reaction, enriched by streptavidin beads, released by selective alkaline hydrolysis of phenacyl ester bond, and subsequently sequenced by LC-MS/MS. This strategy has helped identifying 1115 unique OP adduction sites on 163 proteins in human plasma, and covers lots of OP adducts that cannot be achieved by traditional detection methods. The comprehensive coverage of novel OP substrates provided a general and sensitive approach to retrospective verification and/or dose assessment of toxic OPs.

Epidemiological Surveillance and Mutational Pattern Analysis of Foot-and-Mouth Disease Outbreaks in Bangladesh during 2012–2021

Foot-and-mouth disease (FMD) in cloven-hoofed animals is considered an economically devastating disease in endemic countries like Bangladesh, where the livestock sector contributes to a greater portion of the nation’s economy. The causative agent of the disease, foot-and-mouth disease virus (FMDV), equipped with higher mutational frequency challenges the efficacy of the existing vaccine and control measures. This study, including 32 districts and 71 outbreaks to reveal epidemiological patterns and mutational trends of FMDV over the past 10 years (2012–2021), reported a 54.7% prevalence of FMD, with the majority of outbreaks occurring during the rainy season. Different risk factors such as age, gender, farming system, and vaccination status demonstrated a significant association with FMD cases which was confirmed by the χ2 test ( p &lt; 0.05 ). VP1 sequence analyses reported the predominance of serotype O (85%) over serotype A (11%) and serotype Asia 1 (4%). Bangladesh has foreseen the emergence of several novel FMDV strains during this decade. Novel sublineages, Ind-2001BD1 (Ind-2001e) and Ind-2001BD2, were reported under serotype O, the G-IX lineage of serotype Asia 1 emerged in 2018, and most recently in 2021, a new genotype named MYMBD21 under the lineage SA-2018 was detected for the first time in Bangladesh. Until now, Ind-2001e (Ind-2001BD1) sublineage under serotype O became the predominant sublineage in Bangladesh. From the mutational trend analysis, highly variable sites were observed at positions 138 and 140 within the G-H loop for serotype O. For serotype A and Asia 1, 45th and 44th residues within the B-C loop showed the highest amino acid variations, respectively. A changing mutational pattern among the 2019–2021 FMDV O and A isolates was also observed. The findings of the study would be crucial to understand the FMD situation and designing necessary preventive steps according to the progressive control pathway for FMD control in Bangladesh.

A comprehensive analysis of translational misdecoding pattern and its implication on genetic code evolution.

The universal genetic code is comprised of 61 sense codons, which are assigned to 20 canonical amino acids. However, the evolutionary basis for the highly conserved mapping between amino acids and their codons remains incompletely understood. A possible selective pressure of evolution would be minimization of deleterious effects caused by misdecoding. Here we comprehensively analyzed the misdecoding pattern of 61 codons against 19 noncognate amino acids where an arbitrary amino acid was omitted, and revealed the following two rules. (i) If the second codon base is U or C, misdecoding is frequently induced by mismatches at the first and/or third base, where any mismatches are widely tolerated; whereas misdecoding with the second-base mismatch is promoted by only U-G or C-A pair formation. (ii) If the second codon base is A or G, misdecoding is promoted by only G-U or U-G pair formation at the first or second position. In addition, evaluation of functional/structural diversities of amino acids revealed that less diverse amino acid sets are assigned at codons that induce more frequent misdecoding, and vice versa, so as to minimize deleterious effects of misdecoding in the modern genetic code.

Genetic architecture of tuber-bound free amino acids in potato and effect of growing environment on the amino acid content

Free amino acids in potato tubers contribute to their nutritional value and processing quality. Exploring the natural variation in their accumulation in tubers across diverse genetic backgrounds is critical to potato breeding programs aiming to enhance or partition their distribution effectively. This study assessed variation in the tuber-bound free amino acids in a diversity panel of tetraploid potato clones developed and maintained by the Texas A&M Potato Breeding Program to explore their genetic basis and to obtain genomic-estimated breeding values for applied breeding purposes. Free amino acids content was evaluated in tubers of 217 tetraploid potato clones collected from Dalhart, Texas in 2019 and 2020, and Springlake, Texas in 2020. Most tuber amino acids were not affected by growing location, except histidine and proline, which were significantly lower (− 59.0%) and higher (+ 129.0%), respectively, at Springlake, Texas (a location that regularly suffers from abiotic stresses, mainly high-temperature stress). Single nucleotide polymorphism markers were used for genome-wide association studies and genomic selection of clones based on amino acid content. Most amino acids showed significant variations among potato clones and moderate to high heritabilities. Principal component analysis separated fresh from processing potato market classes based on amino acids distribution patterns. Genome-wide association studies discovered 33 QTL associated with 13 free amino acids. Genomic-estimated breeding values were calculated and are recommended for practical potato breeding applications to select parents and advance clones with the desired free amino acid content.

Mitonuclear interactions shape both direct and parental effects of diet on fitness and involve a SNP in mitoribosomal 16s rRNA.

Nutrition is a primary determinant of health, but responses to nutrition vary with genotype. Epistasis between mitochondrial and nuclear genomes may cause some of this variation, but which mitochondrial loci and nutrients participate in complex gene-by-gene-by-diet interactions? Furthermore, it remains unknown whether mitonuclear epistasis is involved only in the immediate responses to changes in diet, or whether mitonuclear genotype might modulate sensitivity to variation in parental nutrition, to shape intergenerational fitness responses. Here, in Drosophila melanogaster, we show that mitonuclear epistasis shapes fitness responses to variation in dietary lipids and amino acids. We also show that mitonuclear genotype modulates the parental effect of dietary lipid and amino acid variation on offspring fitness. Effect sizes for the interactions between diet, mitogenotype, and nucleogenotype were equal to or greater than the main effect of diet for some traits, suggesting that dietary impacts cannot be understood without first accounting for these interactions. Associating phenotype to mtDNA variation in a subset of populations implicated a C/T polymorphism in mt:lrRNA, which encodes the 16S rRNA of the mitochondrial ribosome. This association suggests that directionally different responses to dietary changes can result from variants on mtDNA that do not change protein coding sequence, dependent on epistatic interactions with variation in the nuclear genome.

Identification of rare nsSNPs in fragile histidine triad (FHIT) gene to explore its correlation with oral cancer: An in-silico approach

Genetic alterations in fragile histidine triad (FHIT) play a pivotal role in various cancers of head, neck, lung, kidney, gastrointestinal, and breast including oral which leads to abnormal transcripts in vivo during the development of oral cancer. Owing to the importance of FHIT gene in cell proliferation, tumor suppression and survival, the structural, functional, non-synonymous SNPs (nsSNPs), and network study was conducted to look at the potential relationship between phenotypic variations and genetic variations. In silico genomic analysis of FHIT was initiated with the identification of 18 rare variants from dbSNP database followed by PredictSNP 1.0 web server analysis for predicting the deleterious and neutral mutants. A total of 11 mutations i.e. P33T, P33A, V97F, S22A, T19I, T61M, D57N, P101A, P101S, S81P and R46H corresponding to 9 nsSNPs were found to be deleterious which affects the protein. The interacting genes with FHIT were found by analyzing protein–protein interactions network using a STRING database. Gene ontology and Disease Association functional analysis of FHIT was obtained by WebGestalt. The mutational positions and amino acid variations have been mapped onto native FHIT. Structural and docking analysis of native and mutant FHIT with ER 27319 Maleate was performed using Auto Dock 4.2, GLIDE, SRide server, structural visualizers, and MD simulations to check their binding energies, stabilizing residues, and to investigate their dynamic behavior, mode of binding action and inhibitor specificity. Our in-silico analysis suggested that screening of P101S (rs533270218), S81P (rs536941406) and D57N (rs375883257) variants of FHIT could be useful for molecular diagnosis and development of vital molecular inhibitors of FHIT pathways. The diagnostic and prognostic approach of these molecular biomarkers can intrude on the predisposition to oral cancer.

Differential Aluminum tolerance in Arabidopsis thaliana ecotypes is seemingly related to metabolite changes

Plants are negatively affected by aluminum (Al) in acidic soils (pH ≤ 5.0), which impairs root growth and ultimately plant yield. Organic acids are closely related to Al neutralization improving the metal tolerance with an expansive metabolic cost. Here, we investigated phenotypic, metabolic, and genetic responses of three Arabidopsis thaliana ecotypes, Columbia (Col-0), Wassilewskija (Ws) and Landsberg erecta (Ler) in response to Al. By comparing the respective control plants with Al-exposed plants, the ecotypes Col-0 and Ws displayed stronger reductions in root growth and reproductive yield than Ler. All ecotypes presented high expression of ALMT1, an Al-resistance associated gene. Further analyses revealed a large accumulation of diverse amino acids, carbohydrates and organic acids following Al stress in both shoot and root tissues of Col-0 and Ws plants, but not in Ler. Altogether, our results suggest that a higher capacity for using and translocating reduced carbon molecules seems crucial to overcoming Al stress in the Ler ecotype. In addition, lower expenses in carbon molecules might be linked to a higher capacity to deal with Al stress, supporting a tight relationship between primary metabolism and Al stress responses. Finally, novel insights on the influence of Al over plant growth and primary metabolism in both shoots and roots are described.

Evolutionary history of MEK1 illuminates the nature of deleterious mutations

Mutations in signal transduction pathways lead to various diseases including cancers. MEK1 kinase, encoded by the human MAP2K1 gene, is one of the central components of the MAPK pathway and more than a hundred somatic mutations in the MAP2K1 gene were identified in various tumors. Germline mutations deregulating MEK1 also lead to congenital abnormalities, such as the cardiofaciocutaneous syndrome and arteriovenous malformation. Evaluating variants associated with a disease is a challenge, and computational genomic approaches aid in this process. Establishing evolutionary history of a gene improves computational prediction of disease-causing mutations; however, the evolutionary history of MEK1 is not well understood. Here, by revealing a precise evolutionary history of MEK1, we construct a well-defined dataset of MEK1 metazoan orthologs, which provides sufficient depth to distinguish between conserved and variable amino acid positions. We matched known and predicted disease-causing and benign mutations to evolutionary changes observed in corresponding amino acid positions and found that all known and many suspected disease-causing mutations are evolutionarily intolerable. We selected several variants that cannot be unambiguously assessed by automated prediction tools but that are confidently identified as "damaging" by our approach, for experimental validation in Drosophila. In all cases, evolutionary intolerant variants caused increased mortality and severe defects in fruit fly embryos confirming their damaging nature. We anticipate that our analysis will serve as a blueprint to help evaluate known and novel missense variants in MEK1 and that our approach will contribute to improving automated tools for disease-associated variant interpretation.

Pharmacology and phytochemical profile of Wattakaka volubilis (L.f.) Stapf: A systematic review

Wattakaka volubilis (L.f.) Stapf, a substantial woody climber of the family Apocynaceae, is widely employed in numerous Ayurvedic formulations to treat leucoderma, asthma, tumors, urinary tract infections, piles, and inflammations. Traditionally, its leaves and entire plant are utilized to alleviate cough, severe cold, fever, rheumatic pain, rabies, snakebites, boils, abscesses, and ocular afflictions. Diverse chemical constituents, encompassing alkaloids, terpenes, amino acids, tannins, pregnane glycosides, flavonoids, and polyphenols, are present in the extracts from different plant parts such as bark, leaves, flowers, and fruits. These extracts exhibit a wide array of pharmacological properties, including anticancer, larvicidal, anti-leishmanial, chondroprotective, and anthelmintic activities. However, for thorough validation, well-designed in vivo and clinical trials are imperative to substantiate its immense medicinal potential. The current investigation aims to deliver a comprehensive review of the bioactive compounds and pharmacological attributes of this plant.

Evolutionarily conserved heat shock protein, HtpX, as an adjunct target against antibiotic-resistant Neisseria gonorrhoeae.

The emergence of multiple drug resistance and extreme drug resistance pathogens necessitates the continuous evaluation of the pathogenic genome to identify conserved molecular targets and their respective inhibitors. In this study, we mapped the global mutational landscape of Neisseria gonorrhoeae (an intracellular pathogen notoriously known to cause the sexually transmitted disease gonorrhoea). We identified highly variable amino acid positions in the antibiotic target genes like the penA, ponA, 23s rRNA, rpoB, gyrA, parC, mtrR and porB. Some variations are directly reported to confer resistance to the currently used front-line drugs like ceftriaxone, cefixime, azithromycinand ciprofloxacin. Further, by whole genome comparison and Shannon entropy analysis, we identified a completely conserved protein HtpX in the drug-resistant as well as susceptible isolates of N. gonorrhoeae (NgHtpX). Comparison with the only available information of Escherichia coli HtpX suggested it to be a transmembrane metalloprotease having a role in stress response. The critical zinc-binding residue of NgHtpX was mapped to E141. By applying composite high throughput screening followed by MD simulations, we identified pemirolast and thalidomide as high-energy binding ligands of NgHtpX. Following cloning and expression of the purified metal-binding domain of NgHtpX (NgHtpXd), its Zn2+ -binding (Kd = 0.4 µM) and drug-binding (pemirolast, Kd = 3.47 µM; and thalidomide, Kd = 1.04 µM) potentials were determined using in-vitro fluorescence quenching experiment. When tested on N. gonorrhoeae cultures, both the ligands imposed a dose-dependent reduction in viability. Overall, our results provide high entropy positions in the targets of presently used antibiotics, which can be further explored to understand the AMR mechanism. Additionally, HtpX and its specific inhibitors identified can be utilised effectively in managing gonococcal infections.