MOESM1 of Improving the thermostability of a fungal GH11 xylanase via site-directed mutagenesis guided by sequence and structural analysis

Additional file 1: Figure S1. SDS-PAGE analysis of the recombinant xylanases. Lanes 1, 3, 5 and 7 correspond to purified XynCDBFV, N207A, G208S, and A210S from E. coli BL21 (DE3), respectively; lanes 2, 4, 6 and 8 correspond to expressed XynCDBFV, N207A, G208S, and A210S, respectively; lane 9 corresponds to control cell (harboring empty pEasy-E2 vector); lane M corresponds to standard protein molecular mass markers

Large Volume Direct Injection Ultra-High Performance Liquid Chromatography–Tandem Mass Spectrometry-Based Comparative Pharmacokinetic Study between Single and Combinatory Uses of Carthamus tinctorius Extract and Notoginseng Total Saponins

The combination of Carthamus tinctorius extract (CTE) and notoginseng total saponins (NGTS), namely, CNP, presents a synergistic effect on myocardial ischemia protection. Herein, comparative pharmacokinetic studies between CNP and CTE/NGTS were conducted to clarify their synergistic mechanisms. A large volume direct injection ultra-high performance liquid chromatography–tandem mass spectrometry (LVDI-UHPLC-MS/MS) platform was developed for sensitively assaying the multi-component pharmacokinetic and in vitro cocktail assay of cytochrome p450 (CYP450) before and after compatibility of CTE and NGTS. The pharmacokinetic profiles of six predominantly efficacious components of CNP, including hydroxysafflor yellow A (HSYA); ginsenosides Rg1 (GRg1), Re (GRe), Rb1 (GRb1), and Rd (GRd); and notoginsenoside R1 (NGR1), were obtained, and the results disclosed that CNP could increase the exposure levels of HSYA, GRg1, GRe, GRb1, and NGR1 at varying degrees. The in vitro cocktail assay demonstrated that CNP exhibited more potent inhibition on CYP1A2 than CTE and NGTS, and GRg1, GRb1, GRd, quercetin, kaempferol, and 6-hydroxykaempferol were found to be the major inhibitory compounds. The developed pharmacokinetic interaction-based strategy provides a viable orientation for the compatibility investigation of herb medicines

Metagenomic Analysis of the Pygmy Loris Fecal Microbiome Reveals Unique Functional Capacity Related to Metabolism of Aromatic Compounds

<div><p>The animal gastrointestinal tract contains a complex community of microbes, whose composition ultimately reflects the co-evolution of microorganisms with their animal host. An analysis of 78,619 pyrosequencing reads generated from pygmy loris fecal DNA extracts was performed to help better understand the microbial diversity and functional capacity of the pygmy loris gut microbiome. The taxonomic analysis of the metagenomic reads indicated that pygmy loris fecal microbiomes were dominated by Bacteroidetes and Proteobacteria phyla. The hierarchical clustering of several gastrointestinal metagenomes demonstrated the similarities of the microbial community structures of pygmy loris and mouse gut systems despite their differences in functional capacity. The comparative analysis of function classification revealed that the metagenome of the pygmy loris was characterized by an overrepresentation of those sequences involved in aromatic compound metabolism compared with humans and other animals. The key enzymes related to the benzoate degradation pathway were identified based on the Kyoto Encyclopedia of Genes and Genomes pathway assignment. These results would contribute to the limited body of primate metagenome studies and provide a framework for comparative metagenomic analysis between human and non-human primates, as well as a comparative understanding of the evolution of humans and their microbiome. However, future studies on the metagenome sequencing of pygmy loris and other prosimians regarding the effects of age, genetics, and environment on the composition and activity of the metagenomes are required.</p> </div

Functional composition of the pygmy loris microbiome.

<p>The percentage of the pygmy loris fecal metagenomic sequences assigned to the general SEED subsystems is shown. Through the “Functional Abundance” tool in MG-RAST, the pygmy loris fecal sequencing runs were determined from the SEED database with the BLASTx algorithm. The e-value cutoff for the metagenomic sequence matches to the SEED subsystem database was 1×10⁻⁵ with a minimum alignment length of 30 bp.</p

Metabolic clustering of pygmy loris, human, mouse, canine, cow, and chicken gastrointestinal metagenomes.

<p>A double hierarchical dendrogram was established through a weight-pair group clustering method based on the non-scaling Manhattan distance. The dendrogram shows the distribution of the functional categories among the ten metagenomes from the six different hosts, including pygmy loris (WFH), humans (HSM and F1S), murine (LMC and OMC), canine (K9C and K9BP), cow (CRP), and chicken (CCA and CCB). The linkages of the dendrogram are based on the relative abundance of metabolic profiles. The heat map depicts the relative percentage of each category of function (variables clustering on the y axis) in each sample (x axis clustering). The heat map color represents the relative percentage of functional categories in each sample, with the legend indicated at the upper left corner. Branch length indicates the Manhattan distances of the samples along the x axis (scale at the upper right corner) and of the microbial classes along the y axis (scale at the lower left corner).</p

Bacterial phylum profiles of the pygmy loris microbiome.

<p>The percentage of the pygmy loris fecal metagenomic sequences assigned to M5NR database is shown. Through the “Organism Abundance” tool in MG-RAST, the pygmy loris fecal sequencing runs were determined from the M5NR database with the BLASTx algorithm. The e-value cutoff for the metagenomic sequence matches to the M5NR database was 1×10⁻⁵, with a minimum alignment length of 30 bp.</p

Phylogenetic clustering of pygmy loris, human, mouse, canine, cow, and chicken gastrointestinal metagenomes.

<p>A double hierarchical dendrogram was established through weight-pair group clustering methods based on the non-scaling Manhattan distance. The dendrogram shows the phylogenetic distribution of the microorganisms among the ten metagenomes from the six different hosts, including pygmy loris (WFH), human (HSM and F1S), mouse (LMC and OMC), dog (K9C and K9BP), cow (CRP), and chicken (CCA and CCB). The linkages of the dendrogram do not show the phylogenetic relationship of the bacterial phylum and are based on the relative abundance of taxonomic profiles. The heat map depicts the relative percentage of each phylum of microorganism (variables clustering on the y axis) in each sample (x axis clustering). The heat map color represents the relative percentage of the microbial descriptions in each sample, with the legend indicated at the upper left corner. Branch length indicates the Manhattan distances of the samples along the x axis (scale at the upper right corner) and of the microbial phyla along the y axis (scale at the lower left corner).</p

Analysis of the protein expressed in <i>E</i>. <i>coli</i> BL21 cells following purification on a 12% SDS-PAGE.

<p>Lane M, protein molecular marker; Lane 1, before induction with 0.7 mM IPTG; Lane 2, after induction with IPTG and grown at 20°C for 20 h; and Lane 3, purified recombinant CarEW (∼53.76 kDa).</p