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Dinoflagellates are protists of high ecological interest due to their ability to produce compounds with biotechnological utility, such as polyunsaturated fatty acids. Two dinoflagellates, Symbiodinium microadriaticum and Perkinsus marinus, play radically different ecological roles. While S. microadriaticum contributes to coral survival by providing nutrients, P. marinus has become a pathogen in oyster culture, causing economic losses. Despite the differences in their ecological role, the mechanism associated which these two organisms have been linked to the production of polyunsaturated fatty acids (PUFAs). Desaturase enzymes play a crucial role in the synthesis of unsaturated fatty acids through the addition of double bonds, especially the Δ4 and Δ5 desaturase enzymes, which are important to producing docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). However, despite their great potential and ecological importance, cellular mechanisms of polyunsaturated fatty acids synthesis and their associated enzymes have not been sufficiently studied in dinoflagellates. This limits efforts to optimizing the production of these metabolites. In this study, an in-silico analysis of Δ4 and Δ5 desaturase sequences from S. microadriaticum and P. marinus was carried out. Bioinformatic characterization showed that genes are related to a membrane-bound proteins with at least 3 transmembrane domains. In addition, three histidine boxes were detected in each enzyme, which are essential for their role as desaturases. In addition, despite distance between protozoan clade and their diverse biological role that S. microadriaticum and P. marinus are involved in, Δ4 and Δ5 desaturases retain high identity domains, even with species from other kingdom such as Plantae and Animalia. [ABSTRACT FROM AUTHOR]

BReast CAncer gene 1 (BRCA1)—a tumor suppressor gene plays an important role in the DNA repair mechanism. Several BRCA1 variants perturb its structure and function, including synonymous and nonsynonymous single nucleotide polymorphisms (SNPs). In the present study, we performed in-silico analyses of nonsynonymous SNPs (nsSNPs) of the BRCA1 gene. In total, 122 nsSNPs were retrieved from the NCBI SNP database and in-silico analyses were performed using computational prediction tools: SIFT, PROVEAN, Mutation Taster, PolyPhen-2, MutPred, and ConSurf. Of these tools, SIFT, PROVEAN, and Mutation Taster predicted 61 out of 122 nsSNPs as "damaging", based on structural homology analysis. PolyPhen-2 classified 22 nsSNPs as "probably damaging". These nsSNPs were further analyzed by MutPred to predict basic molecular mechanisms of amino acid alteration. ConSurf analysis predicted eleven conserved amino acid residues with structural and functional consequences. We identified five amino acid residues in the RING finger domain (L22, C39, H41, C44, and C47) and two in the BRCT domain (P1771 and I1707) with the potential to deter the BRCA1 protein function. This study provides insights into the effect of nsSNPs and amino acid substitutions in BRCA1. [ABSTRACT FROM AUTHOR]

Abstract Background Homozygous inactivating GCK mutations have been repeatedly reported to cause severe hyperglycemia, presenting as permanent neonatal diabetes mellitus (PNDM). Conversely, only two cases of GCK homozygous mutations causing mild hyperglycemia have been so far described. We here report a novel GCK mutation (c.1116G>C, p.E372D), in a family with one homozygous member showing mild hyperglycemia. Methods GCK mutational screening was carried out by Sanger sequencing. Computational analyses to investigate pathogenicity and molecular dynamics (MD) were performed for GCK‐E372D and for previously described homozygous mutations associated with mild (n = 2) or severe (n = 1) hyperglycemia, used as references. Results Of four mildly hyperglycemic family‐members, three were heterozygous and one, diagnosed in the adulthood, was homozygous for GCK‐E372D. Two nondiabetic family members carried no mutations. Fasting glucose (p = 0.016) and HbA1c (p = 0.035) correlated with the number of mutated alleles (0–2). In‐silico predicted pathogenicity was not correlated with the four mutations’ severity. At MD, GCK‐E372D conferred protein structure flexibility intermediate between mild and severe GCK mutations. Conclusions We present the third case of homozygous GCK mutations associated with mild hyperglycemia, rather than PNDM. Our in‐silico analyses support previous evidences suggesting that protein stability plays a role in determining clinical severity of GCK mutations.

Thermotoga maritima derived xylanases are important industrial enzymes owing to extensive thermotolerance and stability under broad pH range This study presents in-silico characterization of Thermotoga maritima derived xylanase A and xylanase B through homology modeling and molecular docking approaches. Analyses of primary and secondary structures revealed net negative charges on both xylanase A and xylanase B. Modular structure of xylanase A exhibited five significant domains including two CBM49, one GlycoHydro10 and two DUF1083. Whereas, structure of xylanase B contained one significant domain i.e. Glyco hydro 10. The amino acid residues Leu-54, Arg-85, Leu-66, Phe-68, Asn-81 and Ile-52 of CBM49 have shown interaction with xylan during docking analyses. Within second CBM49 domain, Tyr-96, Trp-47 and Arg-19 have shown interaction with xylan. No residue of GlycoHydro10 was found to interact with xylan. Whereas, Asn-55, Arg-83, Arg-60 and ILe-118 of DUF1083 interacted with xylan. The Phe-170, Trp-156, Glu-15, Ser-157 and Lys-17 of DUF-1083 domain of xylanase A also showed interaction with xylan. The Glu-15 and Phe-170 of xylanase B showed H-bonding while Lys-565 showed electrostatic interaction with xylan. Comparative phylogenetic analysis of xylanases from Thermotoga genus, showed the complex evolutionary lineage of multimeric xylanase A protein. The findings of this study will lead us towards functional insights of xylanases from Thermotoga maritima and will help the researchers in xylanase enzyme engineering through rational design for developing highly efficient industrial enzymes. [ABSTRACT FROM AUTHOR]

Thalidomide embryopathy (TE) has affected more than 10,000 babies worldwide. The hypothesis of antiangiogenesis as the teratogenic mechanism of thalidomide has been investigated in several experimental models; but, in humans, it has only been accessed by in vitro studies. Here, we hypothesized the effect of thalidomide upon angiogenesis-related molecules or proteins, previously identified in human embryonic cells, through the in silico STRING-tool. We also investigated ten polymorphisms in angiogenesis-related genes in 38 Brazilian TE individuals and 136 non-affected Brazilians. NOS2 , PTGS2 , and VEGFA polymorphisms were chosen for genotyping. The STRING-tool suggested nitric oxide and β-catenin as the central angiogenesis-related molecules affected by thalidomide’s antiangiogenic property. We did not identify a significant difference of allelic, genotypic or haplotypic frequencies between the groups. We could not predict a risk allele or a protective one for TE in NOS2, PTGS2, or VEGFA , although other genes should be analyzed in larger samples. The role of nitric oxide and β-catenin must be further evaluated, regarding thalidomide teratogenesis complex etiology. [ABSTRACT FROM AUTHOR]

β-glucosidases (Bgl) are widely utilized for releasing non-reducing terminal glucosyl residues. Nevertheless, feedback inhibition by glucose end product has limited its application. A noticeable exception has been found for β-glucosidases of the glycoside hydrolase (GH) family 1, which exhibit tolerance and even stimulation by glucose. In this study, using local isolate Trichoderma asperellum UPM1, the gene encoding β-glucosidase from GH family 1, hereafter designated as TaBgl2, was isolated and characterized via in-silico analyses. A comparison of enzyme activity was subsequently made by heterologous expression in Escherichia coli BL21(DE3). The presence of N-terminal signature, cis-peptide bonds, conserved active site motifs, non-proline cis peptide bonds, substrate binding, and a lone conserved stabilizing tryptophan (W) residue confirms the identity of Trichoderma sp. GH family 1 β-glucosidase isolated. Glucose tolerance was suggested by the presence of 14 of 22 known consensus residues, along with corresponding residues L167 and P172, crucial in the retention of the active site's narrow cavity. Retention of 40% of relative hydrolytic activity on ρ-nitrophenyl-β-D-glucopyranoside (ρNPG) in a concentration of 0.2 M glucose was comparable to that of GH family 1 β-glucosidase (Cel1A) from Trichoderma reesei. This research thus underlines the potential in the prediction of enzymatic function, and of industrial importance, glucose tolerance of family 1 β-glucosidases following relevant in-silico analyses. [ABSTRACT FROM AUTHOR]

Background: Class III plant peroxidases play important role in a number of physiological processes in plants such as lignin biosynthesis, suberization, cell wall biosynthesis, reactive oxygen species metabolism and plant defense against pathogens. Peroxidases are also of significance in several industrial applications. In view of this, the production and identification of novel peroxidases having resistance towards temperature, pH, salts is desirable., Objective: The objective of the present work was to clone and characterize a novel plant peroxidase suitable for industrial application., Methods: A full length cDNA clone of lemon peroxidase was isolated using PCR and RACE approaches, characterized and heterologously expressed in Escherichia coli using standard protocols. The expressed peroxidase was purified using Ni-NTA agarose column and biochemically characterized using standard protocols. The peroxidase was also in-silico characterized at nucleotide as well as protein levels using standard protocols., Results: A full length cDNA clone of lemon peroxidase was isolated and expressed heterologously in E. coli. The expressed recombinant lemon peroxidase (LPRX) was activated by in-vitro refolding and purified. The purified LPRX exhibited pH and temperature optima of pH 7.0 and 50°C, respectively. The LPRX was found to be activated by metal ions (Na+, Ca2+, Mg2+ and Mn2+) at lower concentration. The expressional analysis of the transcripts suggested involvement of lemon peroxidase in plant defense. The lemon peroxidase was in silico modelled and docked with the substrates guaiacol, and pyrogallol and shown the favourability of pyrogallol over guaiacol, which is in agreement with the in-vitro findings. The protein function annotation analyses suggested the involvement of lemon peroxidase in the phenylpropanoid biosynthesis pathway and plant defense mechanisms., Conclusion: Based on the biochemical characterization, the purified peroxidase was found to be resistant towards the salts and thus, might be a good candidate for industrial exploitation. The in-silico protein function annotation and transcript analyses highlighted the possible involvement of the lemon peroxidase in plant defense response., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background: Homozygous inactivating GCK mutations have been repeatedly reported to cause severe hyperglycemia, presenting as permanent neonatal diabetes mellitus (PNDM). Conversely, only two cases of GCK homozygous mutations causing mild hyperglycemia have been so far described. We here report a novel GCK mutation (c.1116G>C, p.E372D), in a family with one homozygous member showing mild hyperglycemia. Methods: GCK mutational screening was carried out by Sanger sequencing. Computational analyses to investigate pathogenicity and molecular dynamics (MD) were performed for GCK‐E372D and for previously described homozygous mutations associated with mild (n = 2) or severe (n = 1) hyperglycemia, used as references. Results: Of four mildly hyperglycemic family‐members, three were heterozygous and one, diagnosed in the adulthood, was homozygous for GCK‐E372D. Two nondiabetic family members carried no mutations. Fasting glucose (p = 0.016) and HbA1c (p = 0.035) correlated with the number of mutated alleles (0–2). In‐silico predicted pathogenicity was not correlated with the four mutations' severity. At MD, GCK‐E372D conferred protein structure flexibility intermediate between mild and severe GCK mutations. Conclusions: We present the third case of homozygous GCK mutations associated with mild hyperglycemia, rather than PNDM. Our in‐silico analyses support previous evidences suggesting that protein stability plays a role in determining clinical severity of GCK mutations. [ABSTRACT FROM AUTHOR]

Advancement in bioinformatics with the development of computational tools has enabled the in-silico prediction and identification of transcription regulatory factors and other genetic elements with great ease. In this study, computational analysis of sequence homology of 546 bp 5' region of 16SrRNA gene of Bacillus sp. strain SJ-101 resulted in identification of promoter-like sequences within the rrn gene. Using BPROM tool, the regulatory motifs like -35 and -10 boxes were mapped at 392 and 411 positions, respectively. Furthermore, the cis-acting elements as the binding sites for transcription factors (TF) cpxR and argR were identified at positions 413 and 416 at the upstream of an open reading frame (ORF). The probable functions of the putative TFs were predicted through the Uni-Prot/Swiss-Prot protein database. Search for the Shine-Dalgarno sequence (SD) found the presence of highly conserved SD sequence (AATACC), and a short 42 bp coding sequence/ORF bounded with characteristic transcription start site (AAC) and a stop codon (TGA) at positions 426 and 465 downstream to the promoter elements. A 13 amino acid long translation product of a short ORF has exhibited 100% homology with protein sequences of Bacillus spp., while showing some degree of polymorphism with other reference strains. The comparative homology of the small protein exhibited maximum similarity with Prolyl-4 hydroxylase of Chlamydomonas reinhardtii with 4.11 ZSCORE. The highly conserved regulatory elements and the putative ORF predicted within the 16SrRNA gene may help understand the role of relatively unexplored short ORFs within rrn operon, and their functional products in genetic regulatory mechanisms in eubacteria.