ABSTRACT
Dendrobium loddigesii is a precious traditional Chinese medicine with high medicinal and ornamental value. However, the characterization of its mitochondrial genome is still pending. Here, we assembled the complete mitochondrial genome of D. loddigesii and discovered that its genome possessed a complex multi-chromosome structure. The mitogenome of D. loddigesii consisted of 17 circular subgenomes, ranging in size from 16,323 bp to 56,781 bp. The total length of the mitogenome was 513,356 bp, with a GC content of 43.41%. The mitogenome contained 70 genes, comprising 36 protein-coding genes (PCGs), 31 tRNA genes, and 3 rRNA genes. Furthermore, we detected 403 repeat sequences as well as identified 482 RNA-editing sites and 8154 codons across all PCGs. Following the sequence similarity analysis, 27 fragments exhibiting homology to both the mitogenome and chloroplast genome were discovered, accounting for 9.86% mitogenome of D. loddigesii. Synteny analysis revealed numerous sequence rearrangements in D. loddigesii and the mitogenomes of related species. Phylogenetic analysis strongly supported that D. loddigesii and D. Amplum formed a single clade with 100% bootstrap support. The outcomes will significantly augment the orchid mitochondrial genome database, offering profound insights into Dendrobium's intricate mitochondrial genome architecture.
Subject(s)
Dendrobium , Endangered Species , Genome, Mitochondrial , Phylogeny , Dendrobium/genetics , Dendrobium/classification , Genome, Mitochondrial/genetics , China , RNA, Transfer/genetics , Whole Genome Sequencing , Base Composition , Chromosomes, Plant/genetics , Genome, Plant , Genome, ChloroplastABSTRACT
During the investigations of macrofungi resources in Zhejiang Province, China, an interesting wood rot fungus was collected. Based on morphological and molecular phylogenetic studies, it is described as a new species, Anthracophyllum sinense. A. sinense is characterized by its sessile, charcoal black and pleurotoid pileus, sparse lamellae occasionally branching, clavate basidia with long sterigmata [(3-)6-7(-8) µm], and non-heteromorphous cystidia. A. sinense establishes a separate lineage close to A. archeri and A. lateritium in the phylogenetic tree.
Subject(s)
Agaricales , Basidiomycota , Phylogeny , DNA, Fungal/genetics , ChinaABSTRACT
Research on novel bioactive lipids has garnered increasing interest. Although lipids can be identified by searching mass spectral libraries, the discovery of novel lipids remains challenging as the query spectra of such lipids are not included in libraries. In this study, we propose a strategy to discover novel carboxylic acid-containing acyl lipids by integrating molecular networking with an extended in silico spectral library. Derivatization was performed to improve the response of this method. The tandem mass spectrometry spectra enriched by derivatization facilitated the formation of molecular networking and 244 nodes were annotated. We constructed consensus spectra for these annotations based on molecular networking and developed an extended in silico spectral library based on these consensus spectra. The spectral library included 6879 in silico molecules covering 12,179 spectra. Using this integration strategy, 653 acyl lipids were discovered. Among these, O-acyl lactic acids and N-lactoyl amino acid-conjugated lipids were annotated as novel acyl lipids. Compared with conventional methods, our proposed method allows for the discovery of novel acyl lipids, and extended in silico libraries significantly increase the size of the spectral library.
Subject(s)
Amino Acids , Software , Tandem Mass Spectrometry/methods , Gene Library , Lipids/analysisABSTRACT
The accurate and timely detection of bacteria is critically important for human health as it helps to determine the original source of bacterial infections and prevent disease spread. Herein, gold nanoparticles (AuNPs) were synthesized using polyoxometalates (POMs) as the stabilizing agent. Since AuNPs have glucose oxidase (GOx)-like activity and POMs possess peroxidase (HRP)-like activity, the as-prepared Au@POM nanoparticles have double enzyme-like activities and facilitate cascade reaction. As known, glucose is required as an energy resource during bacterial metabolism, the concentration of glucose decreases with the increase of bacteria content in a system with bacteria and glucose. Therefore, when we use Au@POM nanozymes to trigger the cascade catalysis of glucose and 3,3',5,5'-tetramethylbenzidine (TMB), the concentration of glucose and bacteria can be sensitively detected using the absorbance intensity at 652 nm in the visible spectrum. As demonstration, S. aureus and E. coli were used as model bacteria. The experimental results show that the present method has a good linear relationship in the bacterial concentration range of 1 to 7.5 × 107 colony-forming units (CFU) mL-1 with a detection limit of 5 CFU mL-1. This study shows a great promise of nanozyme cascade reactions in the construction of biosensors and clinical detections.
Subject(s)
Gold , Metal Nanoparticles , Humans , Escherichia coli , Staphylococcus aureus , Glucose , CatalysisABSTRACT
In mass analysis of proteins, mass spectrometry directly measures the mass to charge ratios of ionized proteins and promises higher accuracy than that of indirect approaches measuring other physicochemical properties, provided that the charge states of detected ions are determined. Accurate mass determination of heterogeneously glycosylated proteins is often hindered by unreliable charge determination due to the insufficient resolution of signals from different charge states and inconsistency among mass profiles of ions in individual charge states. Limited charge reduction of a subpopulation of proteoforms using electron transfer/capture reactions (ETnoD/ETnoD) solves this problem by narrowing the mass distribution of examined proteoforms and preserving the mass profile of the precursor charge state in the reduced charge states. However, the limited availability of ETnoD/ETnoD function in commercial instruments limits the application of this approach. Here, utilizing a range of charge-dependent and accuracy-affecting spectral features revealed by a systematic evaluation at levels of both the ensemble and subpopulation of proteoforms based on theoretical models and experiments, we developed a limited charge reduction workflow that enables using collision-induced dissociation and higher energy collisional dissociation, two widely available reactions, as alternatives to ETnoD/ETnoD while providing adequate accuracy. Alternatively, substituting proton transfer charge reduction for ETnoD/ETnoD provides higher accuracy of mass determination. Performing mass selection in a window-sliding manner improves the accuracy and allows profiling of the whole proteoform distribution. The proposed workflow may facilitate the development of universal characterization strategies for more complex and heterogeneous protein systems.
Subject(s)
Proteins , Protons , Electrons , Ions/chemistry , Mass Spectrometry/methods , Proteins/chemistryABSTRACT
RATIONALE: The profiling of natural urinary peptides is a valuable indicator of kidney condition. While front-end separation limits the speed of peptidomic profiling, MS1-based results suffer from limited peptide coverage and specificity. Clinical studies on chronic kidney disease require an effective strategy to balance the trade-off between identification depth and throughput. METHODS: CKD273, a urinary proteome classifier associated with chronic kidney disease, in samples from diabetic nephropathy patients was profiled in parallel using capillary electrophoresis-mass spectrometry (CE-MS), liquid chromatography with mass spectrometry (LC-MS), and matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS). Through cross-comparison of results from MS1 of unfractionated peptides and elution-time-resolved MS1 as well as MS/MS in LC- and CE-MS approaches, we evaluated the contribution of false-positive identification to MS1-based identification and quantitation, and analyzed the benefit of front-end separation in terms of accuracy and efficiency. RESULTS: In LC- and CE-MS, although MS1 data resulted in higher number of identifications than MS/MS, elution-time-dependent analysis revealed extensive interference by non-CKD273 peptides, which would contribute up to 50% to quantitation if they are not separated from genuine CKD273 peptides. In the absence of separation, MS1 data resulted in lower numbers of identifications and abundance pattern that significantly deviated from those by liquid chromatography with tandem mass spectrometry (LC-MS/MS) or capillary electrophoresis with tandem mass spectrometry (CE-MS/MS). CE showed higher identification efficiency even when less sample was used or achieved faster separation. CONCLUSIONS: To ensure the reliability of MS1-based urinary peptide profiling, front-end separation should not be omitted, and elution time should be used in addition to intact mass for identification. Including MS/MS in data acquisition does not compromise the speed or identification number, while benefiting data reliability by providing real-time sequence verification.
ABSTRACT
Tandem mass spectrometry-spectra-based annotation in natural products challenges a lot because of ambiguous structural characterization. It still lacks an efficiency method to score and rank the annotation confidence. Herein, we develop a novel approach to rank the annotation confidences of saponins. Annotations were accomplished according to fragmentation patterns. The corresponding diagnostic fragments and their abundances were recorded. Average abundances were taken as a reference spectrum, and the cosine similarity score (CSS) was calculated to measure how well the spectral matched. According to CSS values, statistic description for confidence levels can be effectively provided. Next, the fragment interpretation score (FIS) was proposed to investigate the deviators' characteristic fragmentation. FIS offset the effect from the deviators' unique fragments. Suspicious annotations involving low CSS and high FIS, may derived from the MS2 spectral background interferences or co-elution. Annotations with low CSS and FIS rank as low confidences, as these annotations need more attention. Using this method, novel saccharide sequences, specific fragmentation preferences, undistinguished precursors, even new structures can also be well traced. By proposed new scoring system, confidence evaluations can be ranked, resulting in significantly enhanced annotation reliability.
Subject(s)
Biological Products , Saponins , Reproducibility of Results , Tandem Mass SpectrometryABSTRACT
The coronavirus pandemic is the biggest in the past 100 years, affected over 200 countries and killed over 300 thousand people. To better understand the epidemics in different areas, the progress percentage was generated in this study by dividing everyday total confirmed case number by the up-to-date total case number, so data obtained from different countries and territories can be put together and compared directly regardless of the large difference in the magnitude of numbers. The global outbreak data were analyzed and categorized into 4 groups based on different epidemic curve stages. The grouping pattern suggests that the geographical position may not play a critical role in the progress of COVID-19 epidemic. In this report, we also used a mathematic model to predict the progress of COVID-19 outbreak in UK, USA and Canada in Group 3, providing valuable information for assessing the risk in these countries and the timing of reopening business.
Subject(s)
Coronavirus Infections/epidemiology , Models, Theoretical , Pneumonia, Viral/epidemiology , Betacoronavirus , COVID-19 , Forecasting , Global Health , Humans , Pandemics , SARS-CoV-2ABSTRACT
BACKGROUND: Phaseolus vulgaris (common bean) microsymbionts belonging to the bacterial genera Rhizobium, Bradyrhizobium, and Ensifer (Sinorhizobium) have been isolated across the globe. Individual symbiosis genes (e.g., nodC) of these rhizobia can be different within each genus and among distinct genera. Little information is available about the symbiotic structure of indigenous Rhizobium strains nodulating introduced bean plants or the emergence of a symbiotic ability to associate with bean plants in Bradyrhizobium and Ensifer strains. Here, we sequenced the genomes of 29 representative bean microsymbionts (21 Rhizobium, four Ensifer, and four Bradyrhizobium) and compared them with closely related reference strains to estimate the origins of symbiosis genes among these Chinese bean microsymbionts. RESULTS: Comparative genomics demonstrated horizontal gene transfer exclusively at the plasmid level, leading to expanded diversity of bean-nodulating Rhizobium strains. Analysis of vertically transferred genes uncovered 191 (out of the 2654) single-copy core genes with phylogenies strictly consistent with the taxonomic status of bacterial species, but none were found on symbiosis plasmids. A common symbiotic region was wholly conserved within the Rhizobium genus yet different from those of the other two genera. A single strain of Ensifer and two Bradyrhizobium strains shared similar gene content with soybean microsymbionts in both chromosomes and symbiotic regions. CONCLUSIONS: The 19 native bean Rhizobium microsymbionts were assigned to four defined species and six putative novel species. The symbiosis genes of R. phaseoli, R. sophoriradicis, and R. esperanzae strains that originated from Mexican bean-nodulating strains were possibly introduced alongside bean seeds. R. anhuiense strains displayed distinct host ranges, indicating transition into bean microsymbionts. Among the six putative novel species exclusive to China, horizontal transfer of symbiosis genes suggested symbiosis with other indigenous legumes and loss of originally symbiotic regions or non-symbionts before the introduction of common bean into China. Genome data for Ensifer and Bradyrhizobium strains indicated symbiotic compatibility between microsymbionts of common bean and other hosts such as soybean.
Subject(s)
Bradyrhizobium/classification , Phaseolus/microbiology , Rhizobium phaseoli/classification , Sinorhizobium/classification , Whole Genome Sequencing/methods , Bradyrhizobium/genetics , Bradyrhizobium/physiology , Chromosomes, Bacterial/genetics , Evolution, Molecular , Gene Transfer, Horizontal , Phylogeny , Plasmids/genetics , Rhizobium phaseoli/genetics , Rhizobium phaseoli/physiology , Root Nodules, Plant/microbiology , Sinorhizobium/genetics , Sinorhizobium/physiology , SymbiosisABSTRACT
The genus Rhizobium usually has a multipartite genome architecture with a chromosome and several plasmids, making these bacteria a perfect candidate for plasmid biology studies. As there are no universally shared genes among typical plasmids, network analyses can complement traditional phylogenetics in a broad-scale study of plasmid evolution. Here, we present an exhaustive analysis of 216 plasmids from 49 complete genomes of Rhizobium by constructing a bipartite network that consists of two classes of nodes, the plasmids and homologous protein families that connect them. Dissection of the network using a hierarchical clustering strategy reveals extensive variety, with 34 homologous plasmid clusters. Four large clusters including one cluster of symbiotic plasmids and two clusters of chromids carrying some truly essential genes are widely distributed among Rhizobium. In contrast, the other clusters are quite small and rare. Symbiotic clusters and rare accessory clusters are exogenetic and do not appear to have co-evolved with the common accessory clusters; the latter ones have a large coding potential and functional complementarity for different lifestyles in Rhizobium. The bipartite network also provides preliminary evidence of Rhizobium plasmid variation and formation including genetic exchange, plasmid fusion and fission, exogenetic plasmid transfer, host plant selection, and environmental adaptation.
Subject(s)
Evolution, Molecular , Plasmids/genetics , Rhizobium/genetics , Phylogeny , Symbiosis/geneticsABSTRACT
Two Gram-stain-negative, rod-shaped bacterial strains (C5T and C16), isolated from root nodules of Phaseolus vulgaris L. in Jiangxi Province, PR China, were characterized by using a polyphasic taxonomical approach. The phylogenetic analysis of the 16S rRNA gene and three concatenated housekeeping genes (recA-glnII-atpD) revealed that C5T and C16 were members of the genus Rhizobium, yet were distinct from known species. The case for strain C5T representing a novel species was supported by genomic results. Pairwise digital DNA-DNA hybridization and average nucleotide identity values were much lower than the proposed and generally accepted species boundaries. The genome-based phylogenetic tree reconstructed by using the up-to-date bacterial core gene set consisting of 92 genes showed that the strains formed a monophyletic branch, further supporting this result. The symbiotic genes of nodC and nifH were identified in both strains; each could nodulate Phaseolus vulgaris and Glycine max but not Leucaena leucocephala, Pisum sativum or Medicago sativa plants. Major cellular fatty acids of C5T were summed feature 8 (C18â:â1 ω7c/C18â:â1 ω6c; 58.8â%), C18â:â1 ω7c 11-methyl (14.2â%) and C18â:â0 (8.1â%). The DNA G+C content of C5T was 61.4 mol%. Based on these genomic, chemotaxonomic and phenotypic characteristics, we propose a novel species: Rhizobium chutanense sp. nov. The type strain is C5T (=CCTCC AB 2018143T=LMG 30777T).
Subject(s)
Phaseolus/microbiology , Phylogeny , Rhizobium/classification , Root Nodules, Plant/microbiology , Bacterial Typing Techniques , Base Composition , China , DNA, Bacterial/genetics , Fatty Acids/chemistry , Genes, Bacterial , Medicago sativa , Nucleic Acid Hybridization , Pisum sativum , RNA, Ribosomal, 16S/genetics , Rhizobium/isolation & purification , Sequence Analysis, DNA , Glycine max , SymbiosisABSTRACT
Dynamic pH junction is an online focusing method in CE based on the electrophoretic mobility difference of analytes in the sample matrix and the background electrolyte. An advantage of this method over the conventional CE is that the sensitivity can be significantly improved. By injecting a long sample plug in the capillary and focusing the analytes at the pH boundary between the background electrolyte and sample matrix, the LOD can be improved by 10-100 folds. The dynamic pH junction method can be easily coupled with ESI-MS. In this work, we used this method for the analysis of microcystins (MCs). The detection limits and dynamic ranges were studied. The separation was optimized by adjusting the injection time, and concentrations and pH values of the background electrolyte. The optimization of analyte focusing leads to enhanced detection response compared to conventional injections, achieving 200-400 fold higher averaged peak heights for four microcystin (MC) variants. More importantly, this method was successfully used for the quantitative analysis of microcystins (MCs) in crude algae samples from natural water bodies, making it promising for practical applications.
Subject(s)
Electrophoresis, Capillary/methods , Mass Spectrometry/methods , Microcystins/analysis , Hydrogen-Ion Concentration , Lakes/chemistry , Limit of Detection , Linear Models , Methanol , Reproducibility of ResultsABSTRACT
Affinity capillary electrophoresis (ACE) in a free solution must be used to confirm the molecular interaction observed in electrospray ionization mass spectrometry (ESI-MS) for affinity binding analysis in the drug discovery process. In this article, the affinity of ibuprofen with three cyclodextrin species of different cavity sizes is investigated by both ESI-MS and ACE methods. Because the binding interactions are measured in different environments using ESI-MS (gas phase) and ACE (liquid phase), the experimental results show significant differences. To better illustrate the major factors influencing the binding constants in different environments, two types of simulations (molecular docking and molecular force field evaluation) are employed in the theoretical discussion. The molecular docking results were consistent with the ESI-MS experimental phenomena. Due to lack of the water molecules in the gas phase, a substantial difference in molecular interaction can contribute to a different binding affinity. By using the molecular modeling based on the force field evaluation with considering solvent molecules, ß-CD is expected to form the most stable complex with ibuprofen in the liquid phase and the binding affinity of ibuprofen to γ-CD is much less significant than it is in gas phase. This prediction is in good agreement with the ACE results. Therefore, this work demonstrates that, by using ESI-MS in the first-step screening and ACE in the follow-up determination, a more accurate understanding can be achieved.
Subject(s)
Electrophoresis, Capillary/methods , Spectrometry, Mass, Electrospray Ionization/methods , Cyclodextrins/chemistry , Molecular Docking Simulation , Solvents/chemistry , WaterABSTRACT
The characterization of alkaloids is challenging because of the diversity of structures and the complicated fragmentation of collision induced structural dissociation in mass spectrometry. In this study, we analyzed the alkaloids in Sinomenium acutum (Thunb.) Rehderet Wil by high resolution mass spectrometry. Chromatographic separation was achieved on a Phenomenex Kinetex C18 (2.1 mm × 100 mm, 2.6 µm) column with a mobile phase consisting of acetonitrile and water (0.1% formic acid) under gradient elution. A total of 52 alkaloids were well separated and 45 of them were structurally characterized, including morphinans, aporphines, benzylisoquinolines, and protoberberines. Specially, mass spectrometric study of the morphinan alkaloids were explicitly investigated. Electrostatic potential plot from simulation was calculated for determination of protonation sites. Further fragmentation analysis suggested that the C3H7N, CH4O, and H2O elimination was displayed in MS² spectrum. These fragmentation pathways are universal for morphinan alkaloids having methoxy substituted cyclohexenone or cyclohexadienone moieties. Additionally, for nitrogen oxides, an ion-neutral complex intermediate is involved in the fragmentation process, generating additional oxygenated ions. All these results provided the universal rules of fragmentation used for detection of alkaloids, and will be expected to be highly useful for comprehensive study of multi-components in the herbal medicine analysis.
Subject(s)
Alkaloids/chemistry , Alkaloids/isolation & purification , Sinomenium/chemistry , Chromatography, Liquid , Mass Spectrometry , Models, Molecular , Molecular Structure , Plant Extracts/chemistry , Plant Extracts/isolation & purificationABSTRACT
Dynamic pH junction focusing prior to electrophoretic separation has been widely used for online pre-concentration of biologically important analytes, which are mostly weakly alkaline/acidic or zwitterionic species such as neurotransmitters, peptides, and proteins. A pH junction is formed when background electrolytes with different pH values are injected sequentially into the separation column of a capillary electrophoresis (CE) system. Unlike the traditional dynamic pH junction configuration with analyte molecules located in a different chemical environment to the separation background electrolyte (BGE), the pH barrage junction has a separate high pH (or low pH) region containing no analyte. Based on Simul 5 Complex simulations and experimental verification with three series of electrolyte combinations, four basic principles for pH barrage junction focusing were identified for its optimization. First, the peak shape after focusing is slightly asymmetric, but this has negligible influence on the analysis result. Second, longer length of the barrage segment is needed for complete focusing with lower concentration of the buffering species. Third, this technique is more advantageous for analytes with relatively high electrophoretic mobility in a capillary without electroosmotic flow. Fourth, provided the analyte region and pH junction buffering species are separated, this quantitative technique is compatible with both optical and mass spectrometric detection.
Subject(s)
Computer Simulation , Electrophoresis, Capillary/methods , Models, Chemical , Hydrogen-Ion Concentration , Mass SpectrometryABSTRACT
Capillary isoelectric focusing directly coupled to high resolution mass spectrometry (cIEF-MS) provides information on amphoteric molecules, including isoelectric point and accurate mass, which enables structural interrogation of biopolymer pI variants. The coupling of cIEF with MS was facilitated by a flow-through microvial interface, made by stainless steel with high chemical resistance and mechanical robustness. Two on-column electrolyte configurations of cIEF-MS were demonstrated using peptide and protein pI markers. The pI resolution was 0.02 pH unit in the pH range of 5.5 to 7.0, with no anticonvective reagent (glycerol) added. High resolution Orbitrap detector provides mass spectra for midsized proteins (<30 kDa), enabling deconvolution with high accuracy for IEF-focused low abundance species. Charge heterogeneity of therapeutic monoclonal antibodies (mAb) is one of the most important attributes in the biopharmaceutical industry, and it is routinely monitored by IEF and fractionation-based methods. As a proof of concept, the commercial formulation of infliximab was directly analyzed using cIEF-MS for separation and online identification of mAb charge variants. The main intact antibody species along with two basic and one acidic variants were observed, and their accurate molecular weights ( Mw) recorded by MS detector readily revealed the structural differences of these variants. Variants with 0.1 unit in pI difference and 1 Da difference in molecular weight were readily resolved. The deconvoluted intact Mw values showed ppm level accuracy compared to theoretical predictions.
Subject(s)
Antibodies, Monoclonal/analysis , Electrophoresis, Capillary/instrumentation , Isoelectric Focusing/instrumentation , Mass Spectrometry/instrumentation , Peptides/analysis , Proteins/analysis , Equipment Design , Glycerol/chemistry , Hydrogen-Ion Concentration , Infliximab/analysis , Isoelectric PointABSTRACT
Recent horizontal gene transfer (HGT) is crucial for enabling microbes to rapidly adapt to their novel environments without relying upon rare beneficial mutations that arise spontaneously. For several years now, computational approaches have been developed to detect HGT, but they typically lack the sensitivity and ability to detect recent HGT events. Here we introduce a novel strategy, named RecentHGT. The number of genes undergoing recent HGT between two bacterial genomes was estimated by a new algorithm derived from the expectation-maximization algorithm and is based on the theoretical sequence-similarity distribution of orthologous genes. We tested the proposed strategy by applying it to a set of 10 Rhizobium genomes, and detected several large-scale recent HGT events. We also found that our strategy was more sensitive than other available HGT detection methods. These HGT events were mainly mediated by symbiotic plasmids. Our new strategy can provide clear evidence of recent HGT events and thus it brings us closer to the goal of detecting these potentially adaptive evolution processes in rhizobia as well as pathogens.
ABSTRACT
This work reveals the deleterious effect of microcystin-LR (MC-LR) on the conformation of DNA and develops an electrochemical biosensor for detection of MC-LR. The biosensor is prepared by physically immobilizing calf thymus DNA (ctDNA) on gold electrode. In the presence of MC-LR, the conformation change of immobilized ctDNA decreases the electron transfer impedance, thus enhances the amperometric response. The proposed method shows a linear range of 4-512â¯ng/L and a detection limit of 1.4â¯ng/L, which is 700-fold lower than the guideline level suggested by the World Health Organization. The detection results are in good agreement with those from the conventional HPLC method. This biosensor possesses good stability against other components in natural water sample, and has been used for detection of MC-LR in local water bodies, indicating its application promise.
Subject(s)
Biosensing Techniques , DNA/chemistry , Electrochemical Techniques , Microcystins/analysis , Electrodes , Gold/chemistry , Marine ToxinsABSTRACT
Native mass spectrometry (MS) is an emerging approach for characterizing biomacromolecular structure and interactions under physiologically relevant conditions. In native MS measurement, intact macromolecules or macromolecular complexes are directly ionized from a non-denaturing solvent, and key noncovalent interactions that hold the complexes together can be preserved for MS analysis in the gas phase. This technique provides unique multi-level structural information such as conformational changes, stoichiometry, topology and dynamics, complementing conventional biophysical techniques. Despite the maturation of native MS and greatly expanded range of applications in recent decades, further dissemination is needed to make the community aware of such a technique. In this review, we attempt to provide an overview of the current body of knowledge regarding major aspects of native MS and explain how such technique contributes to the characterization of biomacromolecular higher-order structure and interactions.
Subject(s)
Spectrometry, Mass, Electrospray Ionization/methods , Macromolecular Substances/chemistry , Macromolecular Substances/metabolism , Molecular Structure , Protein Multimerization , Proteins/chemistry , Proteins/metabolismABSTRACT
Due to the wide cultivation of bean (Phaseolus vulgaris L.), rhizobia associated with this plant have been isolated from many different geographical regions. In order to investigate the species diversity of bean rhizobia, comparative genome sequence analysis was performed in the present study for 69 Rhizobium strains mainly isolated from root nodules of bean and clover (Trifolium spp.). Based on genome average nucleotide identity, digital DNA:DNA hybridization, and phylogenetic analysis of 1,458 single-copy core genes, these strains were classified into 28 clusters, consistent with their species definition based on multilocus sequence analysis (MLSA) of atpD, glnII, and recA. The bean rhizobia were found in 16 defined species and nine putative novel species; in addition, 35 strains previously described as Rhizobium etli, Rhizobium phaseoli, Rhizobium vallis, Rhizobium gallicum, Rhizobium leguminosarum and Rhizobium spp. should be renamed. The phylogenetic patterns of symbiotic genes nodC and nifH were highly host-specific and inconsistent with the genomic phylogeny. Multiple symbiovars (sv.) within the Rhizobium species were found as a common feature: sv. phaseoli, sv. trifolii and sv. viciae in Rhizobium anhuiense; sv. phaseoli and sv. mimosae in Rhizobium sophoriradicis/R. etli/Rhizobium sp. III; sv. phaseoli and sv. trifolii in Rhizobium hidalgonense/Rhizobium acidisoli; sv. phaseoli and sv. viciae in R. leguminosarum/Rhizobium sp. IX; sv. trifolii and sv. viciae in Rhizobium laguerreae. Thus, genomic comparison revealed great species diversity in bean rhizobia, corrected the species definition of some previously misnamed strains, and demonstrated the MLSA a valuable and simple method for defining Rhizobium species.