scholarly journals Rat thimet oligopeptidase: large-scale expression in Escherichia coli and characterization of the recombinant enzyme

1995 ◽  
Vol 309 (1) ◽  
pp. 203-207 ◽  
Author(s):  
N McKie ◽  
P M Dando ◽  
M A Brown ◽  
A J Barrett
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Chemical Properties ◽  
Zinc Content ◽  
Recombinant Enzyme ◽  
Naturally Occurring ◽  
Thimet Oligopeptidase ◽  
Thiol Content

The coding sequence for rat testis thimet oligopeptidase (TOP) (EC 3.4.24.15) was placed under the control of the T7 polymerase/promoter system. Cultures of Escherichia coli transfected with the resulting plasmid expressed the enzyme as a soluble cytoplasmic protein. Medium-scale cultures allowed isolation of the enzyme in quantities of tens of milligrams. The availability of the recombinant enzyme permitted the determination of such chemical properties as epsilon 280 (48,960), zinc content (2 atom/molecule) and available thiol content (8-10/molecule) for TOP. The recombinant enzyme showed the catalytic activities previously reported for the naturally occurring enzyme, so that we can now conclude with confidence that these are all due to TOP and there is no need to postulate the existence of separate ‘Pz-peptidase’ or ‘endo-oligopeptidase A’ enzymes.

scholarly journals Isolation and characterization of pathogenic Escherichia coli bacteriophages from chicken and beef offal

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Celosia Lukman ◽  
Christopher Yonathan ◽  
Stella Magdalena ◽  
Diana Elizabeth Waturangi
Keyword(s):  
Escherichia Coli ◽  
Foodborne Pathogens ◽  
High Efficiency ◽  
Multiplicity Of Infection ◽  
Isolation And Characterization ◽  
Transmission Electron ◽  
Pathogenic Escherichia Coli ◽  
Family Based

Abstract Objective This study was conducted to isolate and characterize lytic bacteriophages for pathogenic Escherichia coli from chicken and beef offal, and analyze their capability as biocontrol for several foodborne pathogens. Methods done in this research are bacteriophage isolation, purification, titer determination, application, determination of host range and minimum multiplicity of infection (miMOI), and bacteriophage morphology. Results Six bacteriophages successfully isolated from chicken and beef offal using EPEC and EHEC as host strain. Bacteriophage titers observed between 109 and 1010 PFU mL−1. CS EPEC and BL EHEC bacteriophage showed high efficiency in reduction of EPEC or EHEC contamination in meat about 99.20% and 99.04%. The lowest miMOI was 0.01 showed by CS EPEC bacteriophage. CI EPEC and BL EPEC bacteriophage suspected as Myoviridae family based on its micrograph from Transmission Electron Microscopy (TEM). Refers to their activity, bacteriophages isolated in this study have a great potential to be used as biocontrol against several foodborne pathogens.

Integrated mass spectrometry-based multi-omics for elucidating mechanisms of bacterial virulence

2021 ◽  
Author(s):  
Lok Man ◽  
William P. Klare ◽  
Ashleigh L. Dale ◽  
Joel A. Cain ◽  
Stuart J. Cordwell
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Large Scale ◽  
Lipid A ◽  
Phenotypic Characterization ◽  
Post Translational Modification ◽  
Form Of Life ◽  
Antimicrobial Interventions

Despite being considered the simplest form of life, bacteria remain enigmatic, particularly in light of pathogenesis and evolving antimicrobial resistance. After three decades of genomics, we remain some way from understanding these organisms, and a substantial proportion of genes remain functionally unknown. Methodological advances, principally mass spectrometry (MS), are paving the way for parallel analysis of the proteome, metabolome and lipidome. Each provides a global, complementary assay, in addition to genomics, and the ability to better comprehend how pathogens respond to changes in their internal (e.g. mutation) and external environments consistent with infection-like conditions. Such responses include accessing necessary nutrients for survival in a hostile environment where co-colonizing bacteria and normal flora are acclimated to the prevailing conditions. Multi-omics can be harnessed across temporal and spatial (sub-cellular) dimensions to understand adaptation at the molecular level. Gene deletion libraries, in conjunction with large-scale approaches and evolving bioinformatics integration, will greatly facilitate next-generation vaccines and antimicrobial interventions by highlighting novel targets and pathogen-specific pathways. MS is also central in phenotypic characterization of surface biomolecules such as lipid A, as well as aiding in the determination of protein interactions and complexes. There is increasing evidence that bacteria are capable of widespread post-translational modification, including phosphorylation, glycosylation and acetylation; with each contributing to virulence. This review focuses on the bacterial genotype to phenotype transition and surveys the recent literature showing how the genome can be validated at the proteome, metabolome and lipidome levels to provide an integrated view of organism response to host conditions.

scholarly journals Fucoidan Characterization: Determination of Purity and Physicochemical and Chemical Properties

Marine Drugs ◽  
2020 ◽  
Vol 18 (11) ◽  
pp. 571
Author(s):  
Ahmed Zayed ◽  
Mona El-Aasr ◽  
Abdel-Rahim S. Ibrahim ◽  
Roland Ulber
Keyword(s):  
Structural Characteristics ◽  
Chemical Properties ◽  
Industrial Applications ◽  
Chemical Structures ◽  
Seasonal Factors ◽  
Molecular Masses ◽  
Current Article ◽  
Downstream Processes

Fucoidans are marine sulfated biopolysaccharides that have heterogenous and complicated chemical structures. Various sugar monomers, glycosidic linkages, molecular masses, branching sites, and sulfate ester pattern and content are involved within their backbones. Additionally, sources, downstream processes, and geographical and seasonal factors show potential effects on fucoidan structural characteristics. These characteristics are documented to be highly related to fucoidan potential activities. Therefore, numerous chemical qualitative and quantitative determinations and structural elucidation methods are conducted to characterize fucoidans regarding their physicochemical and chemical features. Characterization of fucoidan polymers is considered a bottleneck for further biological and industrial applications. Consequently, the obtained results may be related to different activities, which could be improved afterward by further functional modifications. The current article highlights the different spectrometric and nonspectrometric methods applied for the characterization of native fucoidans, including degree of purity, sugar monomeric composition, sulfation pattern and content, molecular mass, and glycosidic linkages.

scholarly journals Preclinical Characterization of Naturally Occurring Polyketide Cyclophilin Inhibitors from the Sanglifehrin Family

2011 ◽  
Vol 55 (5) ◽  
pp. 1975-1981 ◽  
Author(s):  
Matthew A. Gregory ◽  
Michael Bobardt ◽  
Susan Obeid ◽  
Udayan Chatterji ◽  
Nigel J. Coates ◽  
...  
Keyword(s):  
Large Scale ◽  
Scale Production ◽  
Subgenomic Replicon ◽  
Isomerase Activity ◽  
Lead Compounds ◽  
Naturally Occurring ◽  
Large Scale Production ◽  
Pharmacokinetic Properties ◽  
Biosynthetic Engineering

ABSTRACTCyclophilin inhibitors currently in clinical trials for hepatitis C virus (HCV) are all analogues of cyclosporine (CsA). Sanglifehrins are a group of naturally occurring cyclophilin binding polyketides that are structurally distinct from the cyclosporines and are produced by a microorganism amenable to biosynthetic engineering for lead optimization and large-scale production by fermentation. Preclinical characterization of the potential utility of this class of compounds for the treatment of HCV revealed that the natural sanglifehrins A to D are all more potent than CsA at disrupting formation of the NS5A-CypA, -CypB, and -CypD complexes and at inhibition of CypA, CypB, and CypD isomerase activity. In particular, sanglifehrin B (SfB) was 30- to 50-fold more potent at inhibiting the isomerase activity of all Cyps tested than CsA and was also shown to be a more potent inhibitor of the 1b subgenomic replicon (50% effective concentrations [EC50s] of 0.070 μM and 0.16 μM in Huh 5-2 and Huh 9-13 cells, respectively). Physicochemical and mouse pharmacokinetic analyses revealed low oral bioavailability (F< 4%) and low solubility (<25 μM), although the half-lives (t1/2) of SfA and SfB in mouse blood after intravenous (i.v.) dosing were long (t1/2> 5 h). These data demonstrate that naturally occurring sanglifehrins are suitable lead compounds for the development of novel analogues that are less immunosuppressive and that have improved metabolism and pharmacokinetic properties.

Identification of a halophilic α-amylase gene from Escherichia coli JM109 and characterization of the recombinant enzyme

2013 ◽  
Vol 35 (7) ◽  
pp. 1061-1065 ◽  
Author(s):  
Yutuo Wei ◽  
Xiaobo Wang ◽  
Jiayuan Liang ◽  
Xue Li ◽  
Liqin Du ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Recombinant Enzyme ◽  
Amylase Gene ◽  
Escherichia Coli Jm109

Expression of the pyranose 2-oxidase from Trametes pubescens in Escherichia coli and characterization of the recombinant enzyme

2005 ◽  
Vol 120 (4) ◽  
pp. 387-395 ◽  
Author(s):  
Helena Marešová ◽  
Branislav Večerek ◽  
Marcela Hradská ◽  
Nathalie Libessart ◽  
Stanislav Bečka ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Recombinant Enzyme

Surface Heat Treatment Design Methodology of Large-Scale Castings

2006 ◽  
Vol 513 ◽  
pp. 61-68
Author(s):  
P. Kula ◽  
T. Liskiewicz ◽  
T. Pacyniak
Keyword(s):  
Heat Treatment ◽  
Induction Heating ◽  
Design Methodology ◽  
Large Scale ◽  
Chemical Properties ◽  
Surface Heat ◽  
Heating And Cooling ◽  
Reliable Design ◽  
Forced Cooling

The process of induction heating followed by forced cooling of large-scale castings is considered. The main goal of this study is to develop a reliable design methodology of surface heat treatment process to provide high quality of the treated component. It is assured by determination of the material constants and convection coefficient during heating and cooling experiments carried out on the specimens made of material adequate to the considered large-scale casting. The main part of the investigations was preceded by wide analyses of the mechanical and chemical properties of the tested specimens. The presented global design methodology of large-scale castings gives a possibility to predict the temperature distribution on the hardened surface during induction heating and forced cooling. This allows to select the most adequate parameters and apply them in a real specific process. The obtained results have been analyzed and discussed.

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