scholarly journals Molecular and biochemical characterization of an endo-β-1,3-glucanase from the pinewood nematode Bursaphelenchus xylophilus acquired by horizontal gene transfer from bacteria

2005 ◽  
Vol 389 (1) ◽  
pp. 117-125 ◽  
Author(s):  
Taisei KIKUCHI ◽  
Hajime SHIBUYA ◽  
John T. JONES
Keyword(s):  
Gene Transfer ◽  
Horizontal Gene Transfer ◽  
Biochemical Characterization ◽  
Glycosyl Hydrolase ◽  
Functional Characterization ◽  
Nematode Species ◽  
Bursaphelenchus Xylophilus ◽  
Pinewood Nematode ◽  
Sequence Comparisons

We report the cloning and functional characterization of an endo-β-1,3-glucanase from the pinewood nematode Bursaphelenchus xylophilus acquired by horizontal gene transfer from bacteria. This is the first gene of this type from any nematode species. We show that a similar cDNA is also present in another closely related species B. mucronatus, but that similar sequences are not present in any other nematode studied to date. The B. xylophilus gene is expressed solely in the oesophageal gland cells of the nematode and the protein is present in the nematode's secretions. The deduced amino acid sequence of the gene is very similar to glycosyl hydrolase family 16 proteins. The recombinant protein, expressed in Escherichia coli, preferentially hydrolysed the β-1,3-glucan laminarin, and had very low levels of activity on β-1,3-1,4-glucan, lichenan and barley β-glucan. Laminarin was degraded in an endoglucanase mode by the enzyme. The optimal temperature and pH for activity of the recombinant enzyme were 65 °C and pH 4.9. The protein is probably important in allowing the nematodes to feed on fungi. Sequence comparisons suggest that the gene encoding the endo-β-1,3-glucanase was acquired by horizontal gene transfer from bacteria. B. xylophilus therefore contains genes that have been acquired by this process from both bacteria and fungi. These findings support the idea that multiple independent horizontal gene transfer events have helped in shaping the evolution of several different life strategies in nematodes.

Characterization of the relationships in the pinewood nematode species complex (PWNSC) (Bursaphelenchus spp.) using a heterologous unc-22 DNA probe from Caenorhabditis elegans

Parasitology ◽  
1991 ◽  
Vol 102 (2) ◽  
pp. 303-308 ◽  
Author(s):  
P. Abad ◽  
S. Tares ◽  
N. Brugier ◽  
G. De Guiran
Keyword(s):  
Species Complex ◽  
Nematode Species ◽  
Bursaphelenchus Xylophilus ◽  
Similar Species ◽  
Pinewood Nematode ◽  
C Elegans ◽  
The World ◽  
Different Populations ◽  
Serious Disease

SUMMARYPine wilt is the most serious disease of native pines in Japan and potentially the most important nematode disease of conifers in the world. The pinewood nematode Bursaphelenchus xylophilus was found to be the causal agent. Difficulties arose with respect to the precise identity of some isolates of B. xylophilus and of similar species B. mucronatus and B. fraudulentus. Restriction enzyme analyses of repetitive DNA revealed bands specific for the species B. xylophilus, B. mucronatus and B. fraudulentus. Hybridization patterns obtained with unc-22 gene of C. elegans, clearly identified B. xylophilus, B. mucronatus and B. fraudulentus as well as the different geographic isolates of these species. Furthermore, it is possible to define the phylogenetic relationships between the different populations constituting the ‘pine wood nematode’ complex.

Development of an efficient PCR-based diagnosis protocol for the identification of the pinewood nematode, Bursaphelenchus xylophilus (Nematoda: Aphelenchoididae)

Nematology ◽  
2004 ◽  
Vol 6 (2) ◽  
pp. 279-285 ◽  
Author(s):  
Jae Soon Kang ◽  
Kwang Sik Choi ◽  
Sang Chul Shin ◽  
Il Sung Moon ◽  
Sang Gil Lee ◽  
...  
Keyword(s):  
Intergenic Spacer ◽  
Nematode Species ◽  
Bursaphelenchus Xylophilus ◽  
5S Rrna ◽  
Rrna Gene ◽  
Pinewood Nematode ◽  
Pine Wood ◽  
5S Rrna Gene ◽  
Primer Sets ◽  
Species Specific

Abstract Pine wood wilt disease caused by the pine wood nematode, Bursaphelenchus xylophilus , has been a serious problem in the southern regions of Korea. Efficient diagnosis of B. xylophilus from infected pine wood specimens is critical for the management of this pest. Traditional microscopic examination often results in an erroneous identification because a closely related non-pathogenic species, B. mucronatus, has a great degree of morphological similarity to B. xylophilus. In an attempt to search for reliable molecular markers for the discrimination of these species, we have cloned the 5S rRNA genomic DNA fragments containing both coding and intergenic spacer (IGS) regions from B. xylophilus and B. mucronatus through a homology-probing PCR strategy. Sequence analyses revealed that coding sequences of the 5S rRNA gene from the two species are almost identical (98.3% homology) but that the IGS sequences differ substantially between the species. Based on the IGS sequence differences (69.7% homology), we designed species-specific primer sets and developed a PCR-based diagnosis protocol for the identification and discrimination of the two nematode species on a molecular basis.

scholarly journals Legionella dresdenensis sp. nov., isolated from river water

2010 ◽  
Vol 60 (11) ◽  
pp. 2557-2562 ◽  
Author(s):  
Paul Christian Lück ◽  
Enno Jacobs ◽  
Isolde Röske ◽  
Ute Schröter-Bobsin ◽  
Roger Dumke ◽  
...  
Keyword(s):  
Type Strain ◽  
Biochemical Characterization ◽  
Novel Species ◽  
Phenotypic Characterization ◽  
Fatty Acid Profiles ◽  
Sequence Comparisons ◽  
River Elbe ◽  
Macrophage Infectivity Potentiator ◽  
Macrophage Infectivity

Legionella-like isolates, strains W03-356T, W03-357 and W03-359, from three independent water samples from the river Elbe, Germany, were analysed by using a polyphasic approach. Morphological and biochemical characterization revealed that they were Gram-negative, aerobic, non-spore-forming bacilli with a cut glass colony appearance that grew only on l-cysteine-supplemented buffered charcoal yeast extract agar. Phylogenetic analysis based on sequence comparisons of the 16S rRNA, macrophage infectivity potentiator (mip), gyrase subunit A (gyrA), ribosomal polymerase B (rpoB) and RNase P (rnpB) genes confirmed that the three isolates were distinct from recognized species of the genus Legionella. Phenotypic characterization of strain W03-356T based on fatty acid profiles confirmed that it was closely related to Legionella rubrilucens ATCC 35304T and Legionella pneumophila ATCC 33152T, but distinct from other species of the genus Legionella. Serotyping of the isolates showed that they were distinct from all recognized species of the genus Legionella. Strains W03-356T, W03-357 and W03-359 are thus considered to represent a novel species of the genus Legionella, for which the name Legionella dresdenensis sp. nov. is proposed. The type strain is W03-356T (=DSM 19488T=NCTC 13409T).

Characterization of the AMP-binding protein gene family in Arabidopsis thaliana: will the real acyl-CoA synthetases please stand up?

2000 ◽  
Vol 28 (6) ◽  
pp. 955-957 ◽  
Author(s):  
J. Shockey ◽  
J. Schnurr ◽  
J. Browse
Keyword(s):  
Protein Gene ◽  
De Novo ◽  
Functional Characterization ◽  
De Novo Synthesis ◽  
Triacylglycerol Composition ◽  
Sequence Comparisons ◽  
Triacylglycerol Metabolism ◽  
Binding Protein Gene

One of the most prominent and important topics in modern agricultural biotechnology is the manipulation of oilseed triacylglycerol composition. Towards this goal, we have sought to identify and characterize acyl-CoA synthetases (ACSs), which play an important role in both de novo synthesis and modification of existing lipids. We have identified and cloned 20 different genes that bear strong sequence homology to known ACSs from other organisms. Through sequence comparisons and functional characterization, we have identified several members of this group that encode ACSs, while the other genes fall into the broader category of genes for AMP-binding proteins (AMPBPs). Distinguishing ACSs from AMPBPs will simplify our efforts to understand the role of ACS in triacylglycerol metabolism.

scholarly journals Structural and functional characterization of the Severe fever with thrombocytopenia syndrome virus L protein

2020 ◽  
Author(s):  
Dominik Vogel ◽  
Sigurdur Rafn Thorkelsson ◽  
Emmanuelle R. J. Quemin ◽  
Kristina Meier ◽  
Tomas Kouba ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Functional Characterization ◽  
Limiting Factor ◽  
Genome Replication ◽  
Human Pathogens ◽  
Emerging Viruses ◽  
Replication Process ◽  
L Protein ◽  
Severe Fever

ABSTRACTThe Bunyavirales order contains several emerging viruses with high epidemic potential, including Severe fever with thrombocytopenia syndrome virus (SFTSV). The lack of medical countermeasures, such as vaccines and antivirals, is a limiting factor for the containment of any virus outbreak. To develop such antivirals a profound understanding of the viral replication process is essential. The L protein of bunyaviruses is a multi-functional and multi-domain protein performing both virus transcription and genome replication and, therefore, would be an ideal drug target. We established expression and purification procedures for the full-length L protein of SFTSV. By combining single-particle electron-cryo microscopy and X-ray crystallography, we obtained 3D models covering ∼70% of the SFTSV L protein in the apo-conformation including the polymerase core region, the endonuclease and the cap-binding domain. We compared this first L structure of the Phenuiviridae family to the structures of La Crosse peribunyavirus L protein and influenza orthomyxovirus polymerase. Together with a comprehensive biochemical characterization of the distinct functions of SFTSV L protein, this work provides a solid framework for future structural and functional studies of L protein-RNA interactions and the development of antiviral strategies against this group of emerging human pathogens.

scholarly journals Prebiotic Properties of Bacillus Coagulans MA 13: Production of Galactoside Hydrolyzing Enzymes and Characterization of the Transglycosylation Properties of a GH42 β-Galactosidase

2021 ◽  
Author(s):  
Martina Aulitto ◽  
Strazzulli Andrea ◽  
Ferdinando Sansone ◽  
Flora Cozzolino ◽  
Maria Monti ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Food Industry ◽  
Biochemical Characterization ◽  
Glycosyl Hydrolase ◽  
Bacillus Coagulans ◽  
Gastrointestinal Diseases ◽  
Mass Spectrometry Analysis ◽  
Human Consumption ◽  
Spectrometry Analysis

Abstract BackgroundThe spore-forming lactic acid bacterium Bacillus coagulans MA-13 has been isolated from canned beans manufacturing and successfully employed for the sustainable production of lactic acid from lignocellulosic biomass. Among lactic acid bacteria, B. coagulans strains are generally recognized as safe (GRAS) for human consumption. Low-cost microbial production of industrially valuable products such as lactic acid and various enzymes devoted to the hydrolysis of oligosaccharides and lactose, is of great importance to the food industry. Specifically, α- and β-galactosidases are attractive for their ability to hydrolyze not-digestible galactosides present in the food matrix as well as in the human gastrointestinal tract.ResultsIn this work we have explored the potential of B. coagulans MA-13 as a source of metabolites and enzymes to improve the digestibility and the nutritional value of food. A combination of mass spectrometry analysis with conventional biochemical approaches has been employed to unveil the intra- and extra- cellular glycosyl hydrolase (GH) repertoire of B. coagulans MA-13 under diverse growth conditions. The highest enzymatic activity was detected on β-1,4 and α-1,6-glycosidic linkages and the enzymes responsible for these activities were unambiguously identified as a β-galactosidase (GH42) and α-galactosidase (GH36), respectively. Whilst the former has been found only in the cytosol, the latter is localized also extracellularly. The export of this enzyme may occur through a not yet identified secretion mechanism, since a typical signal peptide is missing in the α-galactosidase sequence. A full biochemical characterization of the recombinant β-galactosidase has been carried out and the ability of this enzyme to perform homo- and hetero-condensation reactions to produce galacto-oligosaccharides, has been demonstrated. ConclusionsProbiotics which are safe for human use and are capable of producing high levels of both α-galactosidase and β-galactosidase are of great importance to the food industry. In this work we have proven the ability of B. coagulans MA-13 to over-produce these two enzymes that are commonly used for treatment of gastrointestinal diseases. Moreover, B. coagulans MA-13 can be employed for an eco-friendly production of prebiotics from dairy food waste because of the ability of β-galactosidase to synthesize galacto-oligosaccharides from lactose.

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