In silico analysis of the possible crosstalk between O-linked ß-GlcNAcylation and phosphorylation sites of Disabled 1 adaptor protein in vertebrates.

Disabled 1 (Dab1) is an adaptor protein with essential functions regulated by reelin signaling and affects many biological processes in the nervous system, including cell motility, adhesion, cortical development, maturation, and synaptic plasticity. Posttranslational modifications directly guide the fates of cytoplasmic proteins to complete their functions correctly. Reciprocal crosstalk between O-GlcNAcylation and phosphorylation is a dynamic modification in cytoplasmic proteins. It modulates the functions of the proteins by regulating their interactions with other molecules in response to the continuously changeable cell microenvironment. Although Dab1 contains conserved recognition sites for phosphorylation in their N-terminal protein interaction domain, the O-ß-GlcNAcylation and phosphorylation sites of human Dab1 sequence, their reciprocal crosstalk, and potential kinases catalyzing the phosphorylation remain unknown. In this study, we determined potential thirty-seven O-ß-GlcNAcylation and sixty-seven phosphorylation sites. Conserved twenty-one residues of these glycosylated sites were also phosphorylated with various kinases, including ATM, CKI, DNAPK, GSK3, PKC, PKG, RSK, cdc2, cdk5, and p38MAPK. In addition, we analyzed these conserved sites at our constructed two- and three-dimensional structures of human Dab1 protein. Dab1 protein models were frequently composed of coil structures as well as α-helix and ß-strands. Many of these conserved crosstalk sites between O-ß-GlcNAcylation and phosphorylation were localized at the coil region of the protein model. These findings may guide biochemical, genetic, and glyco-biology based on further experiments about the Dab1 signaling process. Understanding these modifications might change the point of view of the Dab1 signaling process and treatment for pathological conditions in neurodegenerative diseases such as Alzheimer's disease.

Determine genetic variations in heat shock factor gene family (HSFs) and study their effect on the functional and structural characterization of protein in Tali goat.

In this study, the effect of genetic variations of four heat shock transcription factor genes (HSF1, HSF2, HSF4, and HSF5) on the 3 D protein structure and function were studied. We defined the breed-specific genetic variations of pooled DNA of Tali goat that differed from the goat reference sequence (CHI2.0). Disordered regions of HSF proteins were predicted using PONDR. Post-translation changes were studied by several predicted online servers. Then, the structure of the order region of proteins was anticipated by using the Swiss model. Tali goat HSF genes contain a total number of 181, 679, 91, and 301 SNPs for HSF1, 2, 4, and 5, respectively. Also, 5 and 3 variants were identified as nsSNPs in the coding region of HSF4 and HSF5, respectively. (r.145A/S), (r.322P/Y), (r.379T/C) in HSF4 and (r.300Q/P), (r.573E/Q) in HSF5 obtained the tolerant and high confidence (SIFT score) for nsSNPs. More than half of these proteins are predicted to be disordered (56, 50, 52, and 50%, respectively for HSF1, 2, 4, and 5). Phosphorylation, acetylation, glycosylation, and Sumoylation sites of HSFs were compared between Tali goat and reference goat. Three residues S145, S263, and S322 of HSF4 in Tali goat were phosphorylation sites, and in HSF5, the reference goat has a phosphorylation site in S593.

Phylogenetic analysis and characterization of arsenic (As) transforming bacterial marker proteins following isolation of As-tolerant indigenous bacteria.

Marker proteins play a significant role in bacterial arsenic (As) transformation. Phylogenetic analysis and three-dimensional (3D) characteristics of As transforming bacterial marker proteins guide the evolutionary origin and As transforming potential of the species. Indeed, As-tolerant bacteria also show a significant level of As transformation. Hence, characterization of As transforming bacterial marker proteins, isolation of As transforming bacteria, and proper integration of the findings may guide to elucidate how bacteria transform As. Therefore, phylogenetic analysis and 3D characterization of As transforming bacterial marker protein following isolation of potential indigenous As-tolerant indigenous bacteria were done to explore the mechanism of bacterial As transformation. Phylogenetic analysis of ten As transforming marker proteins (arsA, arsB, arsC, arsD, arsR, aioA, arrA, aioB, acr1, and acr3) in 20 potential bacterial genomes (except 19 for the acr3) were studied. Some bacterial genomes featured up to five marker proteins, and therefore, 3D characteristics of the marker proteins were analyzed in those genomes having three-to-five marker proteins. In phylogeny, species in close clades represent their phylogenetic resemblances and may have similar functions. P. aeruginosa, E. coli, and K. pneumonia were found to be more effective due to having the highest number (five) of marker proteins. In 3D protein modeling, most of the marker proteins were found to be active. Among 19 indigenous bacterial isolates, multiple isolates showed tolerance up to 50 mM As(III) and 250 mM As(V), which may potentially transform a significant quantities of As. Hence, integration of the results of phylogenetic analysis, 3D protein characteristics, and As tolerance in the bacterial isolates could guide to explore the mechanism of how bacteria transform As at cellular and molecular levels.

In Silico Characterisation of the Late Embryogenesis Abundant (LEA) Protein Families and Their Role in Desiccation Tolerance in Ramonda serbica Panc.

Ramonda serbica Panc. is an ancient resurrection plant able to survive a long desiccation period and recover metabolic functions upon watering. The accumulation of protective late embryogenesis abundant proteins (LEAPs) is a desiccation tolerance hallmark. To propose their role in R. serbica desiccation tolerance, we structurally characterised LEAPs and evaluated LEA gene expression levels in hydrated and desiccated leaves. By integrating de novo transcriptomics and homologues LEAP domains, 318 R. serbica LEAPs were identified and classified according to their conserved motifs and phylogeny. The in silico analysis revealed that hydrophilic LEA4 proteins exhibited an exceptionally high tendency to form amphipathic α-helices. The most abundant, atypical LEA2 group contained more hydrophobic proteins predicted to fold into the defined globular domains. Within the desiccation-upregulated LEA genes, the majority encoded highly disordered DEH1, LEA1, LEA4.2, and LEA4.3 proteins, while the greatest portion of downregulated genes encoded LEA2.3 and LEA2.5 proteins. While dehydrins might chelate metals and bind DNA under water deficit, other intrinsically disordered LEAPs might participate in forming intracellular proteinaceous condensates or adopt amphipathic α-helical conformation, enabling them to stabilise desiccation-sensitive proteins and membranes. This comprehensive LEAPs structural characterisation is essential to understanding their function and regulation during desiccation aiming at crop drought tolerance improvement.

Structural modeling of two plant UDP-dependent sugar-sugar glycosyltransferases reveals a conserved glutamic acid residue that is a hallmark for sugar acceptor recognition.

Glycosylation is one of the common modifications of plant metabolites, playing a major role in the chemical/biological diversity of a wide range of compounds. Plant metabolite glycosylation is catalyzed almost exclusively by glycosyltransferases, mainly by Uridine-diphosphate dependent Glycosyltransferases (UGTs). Several X-ray structures have been determined for primary glycosyltransferases, however, little is known regarding structure-function aspects of sugar-sugar/branch-forming O-linked UGTs (SBGTs) that catalyze the transfer of a sugar from the UDP-sugar donor to an acceptor sugar moiety of a previously glycosylated metabolite substrate. In this study we developed novel insights into the structural basis for SBGT catalytic activity by modelling the 3d-structures of two enzymes; a rhamnosyl-transferase Cs1,6RhaT - that catalyzes rhamnosylation of flavonoid-3-glucosides and flavonoid-7-glucosides and a UGT94D1 - that catalyzes glucosylation of (+)-Sesaminol 2-O-ß-d-glucoside at the C6 of the primary sugar moiety. Based on these structural models and docking studies a glutamate (E290 or E268 in Cs1,6RhaT or UGT94D1, respectively) and a tryptophan (W28 or W15 in Cs1,6RhaT or UGT94D1, respectively) appear to interact with the sugar acceptor and are suggested to be important for the recognition of the sugar-moiety of the acceptor-substrate. Functional analysis of substitution mutants for the glutamate and tryptophan residues in Cs1,6RhaT further support their role in determining sugar-sugar/branch-forming GT specificity. Phylogenetic analysis of the UGT family in plants demonstrates that the glutamic-acid residue is a hallmark of SBGTs that is entirely absent from the corresponding position in primary UGTs.

Identification and characterization of amphibian SLC26A5 using RNA-Seq.

BACKGROUND: Prestin (SLC26A5) is responsible for acute sensitivity and frequency selectivity in the vertebrate auditory system. Limited knowledge of prestin is from experiments using site-directed mutagenesis or domain-swapping techniques after the amino acid residues were identified by comparing the sequence of prestin to those of its paralogs and orthologs. Frog prestin is the only representative in amphibian lineage and the studies of it were quite rare with only one species identified. RESULTS: Here we report a new coding sequence of SLC26A5 for a frog species, Rana catesbeiana (the American bullfrog). In our study, the SLC26A5 gene of Rana has been mapped, sequenced and cloned successively using RNA-Seq. We measured the nonlinear capacitance (NLC) of prestin both in the hair cells of Rana's inner ear and HEK293T cells transfected with this new coding gene. HEK293T cells expressing Rana prestin showed electrophysiological features similar to that of hair cells from its inner ear. Comparative studies of zebrafish, chick, Rana and an ancient frog species showed that chick and zebrafish prestin lacked NLC. Ancient frog's prestin was functionally different from Rana. CONCLUSIONS: We mapped and sequenced the SLC26A5 of the Rana catesbeiana from its inner ear cDNA using RNA-Seq. The Rana SLC26A5 cDNA was 2292 bp long, encoding a polypeptide of 763 amino acid residues, with 40% identity to mammals. This new coding gene could encode a functionally active protein conferring NLC to both frog HCs and the mammalian cell line. While comparing to its orthologs, the amphibian prestin has been evolutionarily changing its function and becomes more advanced than avian and teleost prestin.

On the biological activity of cytokinin free bases and their ribosides.

MAIN CONCLUSION: The free bases of cytokinins are the biologically active forms of the hormone while cytokinin ribosides become active only upon removal of the ribose residue. Cytokinins (CKs) belong to the classical plant hormones. They were discovered more than 65 years ago, but which molecular forms possess genuine CK activity is still matter of debate. Numerous studies support the view that only the free bases are the biologically active molecules. This standpoint has been challenged in a recent review (Nguyen et al. in Planta 254: 45, 2021) proposing that also CK ribosides may have genuine own CK activity. Here we critically discuss the pros and cons of this viewpoint considering the results of biological assays, CK binding studies, 3D structural data of CK-receptor interaction and mutant analyses. It is concluded that all types of study provide clear and convincing evidence only for biological activity of free bases and not ribosides; the latter are rather a transport form of the hormone without their own biological activity.

Severe epidermolysis bullosa simplex phenotype caused by codominant mutations p.Ile377Thr in keratin 14 and p.Gly138Glu in keratin 5.

Epidermolysis bullosa simplex (EBS) is a rare skin disease usually inherited in an autosomal dominant pattern. EBS is resulting from mutations in keratin 5 (KRT5) and keratin 14 (KRT14) genes encoding the keratins 5 and 14 proteins expressed in the keratinocytes of the basal layer of the epidermis. To date, seven pathogenic mutations have been reported to be responsible for EBS in the Canadian population from the province of Quebec: p.Pro25Leu, p.Leu150Pro, p.Met327Thr and p.Arg559X in KRT5; p.Arg125Ser, p.Ile377Thr and p.Ile412Phe in KRT14. Here, we present a novel French-Canadian patient diagnosed with EBS confined to the soles but presenting a severe complication form including blisters, hyperkeratosis, skin erosions and toenail abnormalities. Mutation screening was performed by direct sequencing of the entire coding regions of KRT5 and KRT14 genes and revealed the previously reported missense heterozygous mutation c. 1130T > C in KRT14 (p.Ile377Thr). Furthermore, this patient is carrying a second mutation in KRT5, c.413G > A (p.Gly138Glu), which has been linked to an increased risk of basal cell carcinoma in the literature. We suspect an impact of the p.Gly138Glu variant on the EBS phenotype severity of the studied patient. The pathogenicity and consequences of both genetic variations were simulated by in silico tools.

Structure and Assembly of Clathrin Cages.

The unusual structure of clathrin, combined with its ability to assemble and disassemble rapidly in cells provides a model system for us to learn about the ways in which proteins can contribute mechanically to a functioning cell. In this article, we discuss the structural properties of clathrin cages and the triskelions which assemble to form them. The function of clathrin depends on the structure of these triskelions and the interactions they make both with each other during assembly and with the adaptor protein network that drives coated vesicle formation. The atomic resolution structure of clathrin domains has been revealed by X-ray crystallography while scattering studies have enabled the shape of a triskelion in solution to be deduced. Cryo-electron microscopy maps have shown the secondary structure of entire cages, how individual triskelion legs are arranged to form a cage and enabled some bound adaptor proteins to be located. Cage formation itself is energetically finely balanced and requires specific interactions between triskelion legs to be productive, as biochemical studies and in silico modeling have shown. Theoretical, structural and cell biological investigations over many years have contributed to our knowledge of clathrin structure and assembly. It now remains to determine the precise nature of the interactions which occur between clathrin triskelions, light chain and heavy chain and the adaptor protein network.

Ultrastructural Analysis of Prune DwarfVirus Intercellular Transport and Pathogenesis.

Prune dwarf virus (PDV) is an important viral pathogen of plum, sweet cherry, peach, and many herbaceous test plants. Although PDV has been intensively investigated, mainly in the context of phylogenetic relationship of its genes and proteins, many gaps exist in our knowledge about the mechanism of intercellular transport of this virus. The aim of this work was to investigate alterations in cellular organelles and the cell-to-cell transport of PDV in Cucumis sativus cv. Polan at ultrastructural level. To analyze the role of viral proteins in local transport, double-immunogold assays were applied to localize PDV coat protein (CP) and movement protein (MP). We observe structural changes in chloroplasts, mitochondria, and cellular membranes. We prove that PDV is transported as viral particles via MP-generated tubular structures through plasmodesmata. Moreover, the computer-run 3D modeling reveals structural resemblances between MPs of PDV and of Alfalfa mosaic virus (AMV), implying similarities of transport mechanisms for both viruses.

The expression of hypoxia-inducible factor-1α gene is not affected by low-oxygen conditions in yellow perch (Perca flavescens) juveniles.

Hypoxia can affect various fish populations, including yellow perch Perca flavescens, which is an economically and ecologically important species in Lake Erie, a freshwater system that often experiences hypoxia in the hypolimnetic part of the lake. Fish, similarly to mammals, possess molecular oxygen sensor-hypoxia-inducible factor-1 (HIF-1), a transcription factor that can affect expression of many downstream genes related to animal growth and locomotion, protein synthesis, as well as ATP and amino acid metabolism. HIF-1 is a heterodimer, which consists of two subunits: oxygen-sensitive and oxygen-insensitive subunits, α and ß, respectively. In this study, we report first on the molecular cloning and sequencing of P. flavescens HIF-1α. The full-length complementary DNA (cDNA) was isolated and submitted to the GenBank with accession number KT783483. It consists of 3529 base pairs (bp) carrying a single open-reading frame that encompasses 2250 bp of the coding region, 247 bp of the 5' untranslated region (UTR), and 1032 bp of the 3' UTR. The "de novo" prediction of the 3D structure of HIF-1α protein, which consists of 749 amino acids, is presented, too. We then utilized One-Step Taqman® real-time RT-PCR technology to monitor changes in HIF-1α messenger RNA (mRNA) copies in response to chronic hypoxic stress. An experiment was conducted using 14-day post-swim-up stage yellow perch larvae with uninflated swim bladders. This experiment included three treatment groups: hypoxia, mid-hypoxia, and normoxia, in four replicates (four tanks per treatment) with the following dissolved oxygen levels: 3, 4, and >7 mg O2/L, respectively. At the end (2 weeks) and in the middle (1 week) of the experiment, fish from each tank were sampled for body measurements and molecular biology analysis. The results showed no differences in survival (∼90%) between treatment groups. Oxygen concentration was lowered to 3.02 ± 0.15 (mean ± SE) mg O2/L with no adverse effect on fish survival. The highest growth rate was observed in the normoxic group. A similar trend was observed with fish body length. The growth rate of fish declined with decreasing water-dissolved oxygen. The number of HIF-1α mRNA copies was not significantly different between hypoxic, mid-hypoxic, and normoxic conditions, and this was true for fish obtained in the middle and at the end of the experiment. Graphical abstract.

Comparative and evolutionary analysis of α-amylase gene across monocots and dicots.

α-amylase is an important enzyme involved in starch degradation to provide energy to the germinating seedling. The present study was conducted to reveal structural and functional evolution of this gene among higher plants. Discounting polyploidy, most plant species showed only a single copy of the gene making multiple isoforms in different tissues and developmental stages. Genomic length of the gene ranged from 1472 bp in wheat to 2369 bp in soybean, and the size variation was mainly due to differences in the number and size of introns. In spite of this variation, the intron phase distribution and insertion sites were mostly conserved. The predicted protein size ranged from 414 amino acid (aa) in soybean to 449aa in Brachypodium. Overall, the protein sequence similarity among orthologs ranged from 56.4 to 97.4 %. Key motifs and domains along with their relative distances were conserved among plants although several species, genera, and class specific motifs were identified. The glycosyl hydrolase superfamily domain length varied from 342aa in soybean to 384aa in maize and sorghum while length of the C-terminal ß-sheet domain was highly conserved with 61aa in all monocots and Arabidopsis but was 59aa in soybean and Medicago. Compared to rice, 3D structure of the proteins showed 89.8 to 91.3 % similarity among the monocots and 72.7 to 75.8 % among the dicots. Sequence and relative location of the five key aa required for the ligand binding were highly conserved in all species except rice.

A novel selection signature in stearoyl-coenzyme A desaturase (SCD) gene for enhanced milk fat content in Bubalus bubalis.

Modern molecular interventions are dynamic gears for breeding animals with superior genetic make-up. These scientific efforts lead us toward sustainable dairy herds with improved milk production in terms of yield and quality. Many of candidate genes have been dissected at molecular level, and suitable genetic markers have been identified in cattle, but this work has not been validated in buffaloes so far. Stearoyl-coenzyme A desaturase (SCD) has been a potential candidate gene for fat content of milk. Genomic analysis of SCD revealed a total of six variations that were identified through DNA sequencing of animals with lower and higher butter fat %age. After statistical analysis, genotype AB of p.K158I could be associated (P value <0.0001) with higher milk fat %age (10.5 ± 0.5464). This SNP was validated on larger data set by cleaved amplified polymorphic sequences (CAPS) by using DdeI. To scrutinize the functional consequences of p.K158I, 3D protein structure of SCD was predicted by homology modeling and this variation was found located in the vicinity of functional domain and a part of transmembrane helix of this membrane integrated protein. This is a first report toward genetic screening of SCD gene at molecular level in buffalo. This report illustrates the implication of SCD gene and in particular p.K158I variation, in imparting its effect on milk fat %age, which can be targeted in selection of superior dairy buffaloes.

Production of recombinant proteins including the B-cell epitopes of autolysin A of Staphylococcus aureus isolated from clinical sheep mastitis and their potential for vaccine development.

Staphylococcus aureus is the most common clinical mastitis-associated pathogen in sheep which contributes to reduced welfare of affected animals and, therefore, compromises the quality and quantity of milk production. To prevent mastitis and its spread, it is essential to guarantee adequate breeding conditions and animal health, through the adoption of good farm management practices and the application of suitable biosecurity measures. Vaccination can play a strategic role in prevention, control, and eradication of diseases. The identification of secreted and cellular antigens of the predominant sheep-CC130/ST700/t1773 lineage would assist in the design of effective vaccine against mammary infections caused by S. aureus. In the current study, we carried out a 3D structural prediction analysis with the identification of the best B cell epitopes of the whole and secreted portion of S. aureus AtlA. Fragments of atlA, containing the main predicted epitopes, were amplified, cloned, and expressed in Escherichia coli for recombinant protein production. Two selected clones produced recombinant proteins (rAtl4 and rAtl8) showing strong reactivity with a hyperimmune serum against the native AtlA and with blood sera collected from sheep with clinical S. aureus mastitis. These may represent potential candidate protein-based vaccines able to elicit a protective immune response to be evaluated by vaccination and subsequent challenge of the vaccinated sheep.

A Missense Mutation (c.1037 G > C, p. R346P) in PAPSS2 Gene Results in Autosomal Recessive form of Brachyolmia Type 1 (Hobaek Form) in A Consanguineous Family.

BACKGROUND: Brachyolmia is a skeletal disorder with an autosomal mode of inheritance (both dominant and recessive) in which the patients have a short height, scoliosis and a reduced trunk size. METHODS: From the Muzaffargarh District in Pakistan, a consanguineous family with multiple Brachyolmia-affected subjects were enrolled in the present study. Basic epidemiological data and radiographs were collected for the subjects. Whole exome sequencing (WES) which was followed by Sanger sequencing was applied to report the geneticbasic of Brachyolmia. RESULTS: The WES identified a missense mutation (c.1037 G > C, p. R346P) in exon 9 of the PAPSS2 gene that was confirmed by the Sanger sequencing in the enrolled subjects. The mutation followed a Mendalian pattern with an autosomal recessive inheritance mode. Multiple sequence alignment by Clustal Omega indicated that the PAPSS2 mutation-containing domain is highly conserved. The HEK293T whole-cell extract that was transfected with the Myc-tagged PCMV6-PAPSS2 of both the wild and mutant constructs were resolved by SDS-PAGE as well as by a Western blot, which confirmed that there are different PAPSS2 protein expression patterns when they were compared between the control and Brachyolmia patients. This difference between the normal and mutated protein was not evident when the three-dimensional computational structures were generated using homology modeling. CONCLUSION: We report a missense mutation (c.1037 G > C, p. R346P) in the PAPSS2 gene that caused Brachyolmia in a consanguineous Pakistani family.

Antimicrobial Proteins and Peptides in Avian Eggshell: Structural Diversity and Potential Roles in Biomineralization.

The calcitic avian eggshell provides physical protection for the embryo during its development, but also regulates water and gaseous exchange, and is a calcium source for bone mineralization. The calcified eggshell has been extensively investigated in the chicken. It is characterized by an inventory of more than 900 matrix proteins. In addition to proteins involved in shell mineralization and regulation of its microstructure, the shell also contains numerous antimicrobial proteins and peptides (AMPPs) including lectin-like proteins, Bacterial Permeability Increasing/Lipopolysaccharide Binding Protein/PLUNC family proteins, defensins, antiproteases, and chelators, which contribute to the innate immune protection of the egg. In parallel, some of these proteins are thought to be crucial determinants of the eggshell texture and its resulting mechanical properties. During the progressive solubilization of the inner mineralized eggshell during embryonic development (to provide calcium to the embryo), some antimicrobials may be released simultaneously to reinforce egg defense and protect the egg from contamination by external pathogens, through a weakened eggshell. This review provides a comprehensive overview of the diversity of avian eggshell AMPPs, their three-dimensional structures and their mechanism of antimicrobial activity. The published chicken eggshell proteome databases are integrated for a comprehensive inventory of its AMPPs. Their biochemical features, potential dual function as antimicrobials and as regulators of eggshell biomineralization, and their phylogenetic evolution will be described and discussed with regard to their three-dimensional structural characteristics. Finally, the repertoire of chicken eggshell AMPPs are compared to orthologs identified in other avian and non-avian eggshells. This approach sheds light on the similarities and differences exhibited by AMPPs, depending on bird species, and leads to a better understanding of their sequential or dual role in biomineralization and innate immunity.

Structural aspects of ß-glucosidase of Myceliophthora thermophila (MtBgl3c) by homology modelling and molecular docking.

Cellulases are the enzymes with diverse range of industrial applications. Cellulases degrade cellulose into monomeric glucose units by hydrolysing ß-1,4-glycosidic bonds. There are three components of cellulases: a) endoglucanase, b) exoglucanase and c) ß-glucosidase which act synergistically in cellulose bioconversion. The cellulases are the third largest industrial enzymes with a great potential in bioethanol production. In this investigation, a ß-glucosidase of a thermophilic fungus Myceliophthora thermophila (MtBgl3c) was analysed for its structural characterization using in silico approaches. The protein structure of MtBgl3c is unknown, therefore an attempt has been made to model 3D structure using Modeller 9.23 software. The MtBgl3c protein model generated was validated from Verify 3D and ERRAT scores of 89.37% and 71.25%, respectively derived from SAVES. Using RAMPAGE the Ramachandran plot was generated, which predicted the accuracy of the 3D model with 91.5% amino acid residues in the favored region. The ion binding and N-glycosylation sites were also predicted. The generated model was docked with cellobiose to predict the most favorable binding sites of MtBgl3c. The key amino acid residues involved in cellobiose bonding are Val88, Asp106, Asp287, Tyr255, Arg170, Glu514. The catalytic conserved amino residues of MtBgl3c were identified. The dock score of cellobiose with MtBgl3c is much lower (-6.46 kcal/mol) than that of glucose (-5.61 kcal/mol), suggesting its high affinity for cellobiose. The docking data of MtBgl3c with glucose illustrate its tolerance to glucose. The present study provides insight into structural characteristics of the MtBgl3c which can be further validated by experimental data. Highlights3D structure of ß-glucosidase (MtBgl3c) of Myceliophthora thermophila is being proposed based on computational analysesThe amino acid residues Asp106, Asp287, Tyr255, Arg170 and Glu514 have been identified to play catalytically important role in substrate bindingDocking and interaction of MtBgl3c with cellobiose and glucose has been confirmedDocking analysis of MtBgl3c with glucose suggested its glucose toleranceThe data would be useful in engineering enzymes for attaining higher catalytic efficiencyCommunicated by Ramaswamy H. Sarma.

PDBsum1: A standalone program for generating PDBsum analyses.

PDBsum1 is a standalone set of programs to perform the same structural analyses as provided by the PDBsum web server (https://www.ebi.ac.uk/pdbsum). The server has pages for every entry in the Protein Data Bank (PDB) and can also process user-uploaded PDB files, returning a password-protected set of pages that are retained for around 3 months. The standalone version described here allows for in-house processing and indefinite retention of the results. All data files and images are pre-generated, rather than on-the-fly as in the web version, so can be easily accessed. The program runs on Linux, Windows, and mac operating systems and is freely available for academic use at https://www.ebi.ac.uk/thornton-srv/software/PDBsum1.

Bioinformatics Predicted Linear Epitopes of the Major Coat Protein of the Beet Yellows Virus for Detection of the Virus in the Cell Extract of the Infected Plant.

Beet yellows virus, which belongs to the genus Closterovirus, family Closteroviridae and has a significant negative economic impact, has proven to be challenging to detect and diagnose. To obtain antibodies against BYV, we propose an easier bioinformatics approach than the isolation and purification of the wild virus as an antigen. We used the SWISS-MODEL Workspace (Biozentrum Basel) protein 3D prediction program to discover epitopes of major coat protein p22 lying on the surface of the BYV capsid. Sequences coding these epitopes were cloned into plasmid pQE-40 (Qiagen) in frame with mouse dihydrofolate reductase gene. Fused epitopes were expressed in Escherichia coli and isolated by the Ni-NTA affinity chromatography. Murine antibodies were raised against each epitope and in a combination of both and characterized by dot-ELISA and indirect ELISA. We successively used these antibodies for diagnosis of virus disease in systemically infected Tetragonia tetragonioides. We believe the approach described above can be used for diagnostics of difficult-to-obtain and hazardous-to-health viral infections.

PDBsum extras: SARS-CoV-2 and AlphaFold models.

The PDBsum web server provides structural analyses of the entries in the Protein Data Bank (PDB). Two recent additions are described here. The first is the detailed analysis of the SARS-CoV-2 virus protein structures in the PDB. These include the variants of concern, which are shown both on the sequences and 3D structures of the proteins. The second addition is the inclusion of the available AlphaFold models for human proteins. The pages allow a search of the protein against existing structures in the PDB via the Sequence Annotated by Structure (SAS) server, so one can easily compare the predicted model against experimentally determined structures. The server is freely accessible to all at http://www.ebi.ac.uk/pdbsum.