Improved expression and purification of the correctly folded response regulator PlnC from lactobacilli.

Abstract:

:The response regulator PlnC is part of the signal transduction system that plays a key role in the regulation of bacteriocin production in Lactobacillus plantarum C11. In this study, we wanted to express high levels of the response regulator PlnC in a soluble and native form for purification and further studies. The protein was expressed as a fusion protein (fPlnC) containing an N-terminal Flag-tag to facilitate detection and purification. When the fusion gene, fplnC, was expressed in Escherichia coli BL21, nearly all (99%) of the recombinant protein ended up inside inclusion bodies as an incorrectly folded protein. By utilizing two different Gram-positive expression systems (SIP and NICE) in L. plantarum NC8 and Lactobacillus sakei Lb790, the expression of the soluble fPlnC was significantly increased, being 20-40 times more than that in E. coli BL21. Using the N-terminal tag, the expressed protein was purified by immunoprecipitation. By DNA-binding study (EMSA), we demonstrated that the fusion protein purified from the soluble pool was correctly folded as judged by its ability to bind specifically on regulated promoters. Using our approach, we estimate that about 1 mg of fPlnC can be purified from 11 of the bacterial culture.

journal_name

J Microbiol Methods

authors

Straume D,Axelsson L,Nes IF,Diep DB

doi

10.1016/j.mimet.2006.03.022

subject

Has Abstract

pub_date

2006-11-01 00:00:00

pages

193-201

issue

2

eissn

0167-7012

issn

1872-8359

pii

S0167-7012(06)00094-7

journal_volume

67

pub_type

杂志文章
  • Real-time PCR for quantification of eleven individual Fusarium species in cereals.

    abstract::Contamination of cereals with Fusarium species is one of the major sources of mycotoxins in food and feed. Quantification of biomass of Fusarium species is essential to understand the interactions of individual species in disease development. In this study quantitative real-time PCR assays based on the elongation fact...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2008.10.016

    authors: Nicolaisen M,Suproniene S,Nielsen LK,Lazzaro I,Spliid NH,Justesen AF

    更新日期:2009-03-01 00:00:00

  • A new recombinant protein expression system for high-throughput screening in the yeast Yarrowia lipolytica.

    abstract::Development of a high-throughput eukaryotic screening procedure is important to increase success in obtaining improved enzymes through directed enzyme evolution. This procedure was developed for the yeast Yarrowia lipolytica which becomes the second eukaryotic host for this purpose. The extracellular lipase Lip2 was u...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2007.06.008

    authors: Bordes F,Fudalej F,Dossat V,Nicaud JM,Marty A

    更新日期:2007-09-01 00:00:00

  • Investigations of the relationship between use of in vitro cell culture-quantitative PCR and a mouse-based bioassay for evaluating critical factors affecting the disinfection performance of pulsed UV light for treating Cryptosporidium parvum oocysts in sa

    abstract::Cryptosporidium parvum is an enteric coccidian parasite that is recognised as a frequent cause of water-borne disease in humans. We report for the first time on use of the in vitro HCT-8 cell culture-quantitative PCR (qPCR) assay and the in vivo SCID-mouse bioassay for evaluating critical factors that reduce or elimin...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2010.01.017

    authors: Garvey M,Farrell H,Cormican M,Rowan N

    更新日期:2010-03-01 00:00:00

  • Assessing the performance of novel software Strain Solution on automated discrimination of Escherichia coli serotypes and their mixtures using matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    abstract::O157, O26, and O111 are the most important O serogroups of enterohemorrhagic Escherichia coli worldwide. Recently we reported a strategy for discriminating these serotypes from the others using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) based on the S10-spc-alpha operon...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2015.11.005

    authors: Ojima-Kato T,Yamamoto N,Iijima Y,Tamura H

    更新日期:2015-12-01 00:00:00

  • Analysis of a diverse assemblage of diazotrophic bacteria from Spartina alterniflora using DGGE and clone library screening.

    abstract::Methods to assess the diversity of the diazotroph assemblage in the rhizosphere of the salt marsh cordgrass, Spartina alterniflora were examined. The effectiveness of nifH PCR-denaturing gradient gel electrophoresis (DGGE) was compared to that of nifH clone library analysis. Seventeen DGGE gel bands were sequenced and...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2008.02.005

    authors: Lovell CR,Decker PV,Bagwell CE,Thompson S,Matsui GY

    更新日期:2008-05-01 00:00:00

  • Detection of a whitening fluorescent agent as an indicator of white paper biodegradation: a new approach to study the kinetics of cellulose hydrolysis by mixed cultures.

    abstract::A simple and reliable method to estimate paper degradation by cellulolytic bacteria is described. This method is based on the detection in the culture medium of a fluorescent whitening agent (FWA) added to white paper during the manufacturing process. Preliminary results using a Cellulomonas strain cultivated in a liq...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/s0167-7012(99)00043-3

    authors: Bichet-Hébé I,Pourcher AM,Sutra L,Comel C,Moguedet G

    更新日期:1999-08-01 00:00:00

  • Efficient recovery of low endotoxin medium-chain-length poly([R]-3-hydroxyalkanoate) from bacterial biomass.

    abstract::Bacterial polyhydroxyalkanoate (PHA) is an attractive biopolyester for medical applications due to its biocompatibility. However, inappropriate extraction of PHA from bacterial biomass results in contamination by pyrogenic compounds (e.g. lipopolysaccharides) and thus influences medical testing. This problem was solve...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2007.01.002

    authors: Furrer P,Panke S,Zinn M

    更新日期:2007-04-01 00:00:00

  • Isolation and concentration of Salmonellae with an immunoaffinity column.

    abstract::A method for rapidly and selectively isolating Salmonellae from buffer solutions and concentrating the bacteria by a factor of approximately 500 was developed. Anti-Salmonellae antibody was covalently linked to 40 microm polyacrylamide beads to prepare a solid phase with affinity for the bacteria. The beads were packe...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/s0167-7012(03)00163-5

    authors: Brewster JD

    更新日期:2003-10-01 00:00:00

  • Efficient differentiation of Corynebacterium striatum, Corynebacterium amycolatum and Corynebacterium xerosis clinical isolates by multiplex PCR using novel species-specific primers.

    abstract::A multiplex-PCR (mPCR) assay was designed with species-specific primers which generate amplicons of 226bp, 434bp and 106bp for differentiating the species C. striatum, C. amycolatum, and C. xerosis, respectively. mPCR results were 100% in agreement with identifications achieved by 16S rRNA and rpoB gene sequencing and...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2017.09.002

    authors: Santos CS,Ramos JN,Vieira VV,Pinheiro CS,Meyer R,Alcantara-Neves NM,Ramos RT,Silva A,Hirata R Jr,Felicori L,de Alegría Puig CR,Navas J,Azevedo V,Mattos-Guaraldi AL,Pacheco LGC

    更新日期:2017-11-01 00:00:00

  • Bacillus thuringiensis improved isolation methodology from soil samples.

    abstract::Bacillus thuringiensis is a sporulating bacterium, which produces parasporal inclusions toxic to insects. Widely used methodologies to isolate Bt from soil consist of a thermal shock treatment followed by selective germination of spores. The results presented here suggest that a preliminary 5 h dry-heat treatment larg...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2008.06.008

    authors: Santana MA,Moccia-V CC,Gillis AE

    更新日期:2008-10-01 00:00:00

  • A new chromogenic agar medium for detection of potentially virulent Yersinia enterocolitica.

    abstract::Several outbreaks of foodborne yersiniosis have been documented and this disease continues to be source of infections transmitted through foods. The selective agars most commonly used to isolate Yersinia enterocolitica in clinical, food and environmental samples, cefsulodin-irgasan-novobiocin (CIN) and MacConkey (MAC)...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2007.11.019

    authors: Weagant SD

    更新日期:2008-02-01 00:00:00

  • Epidemiological investigation and glycotyping of clinical Pseudomonas aeruginosa isolates from patients with cystic fibrosis by mass spectrometry: association with multiple drug resistance.

    abstract::The aim of the present investigation was to develop a novel approach for rapid identification and differentiation of Pseudomonas aeruginosa clones from Canadian cystic fibrosis patients by capillary electrophoresis-mass spectrometry. We screened P. aeruginosa isolates for lipopolysaccharide structure and presence/abse...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2008.10.008

    authors: Altman E,Wang Z,Aaron SD,Liu X,Vandemheen KL,Ferris W,Giesbrecht T,Li J

    更新日期:2009-02-01 00:00:00

  • Improved pH-ratiometry for the three-dimensional mapping of pH microenvironments in biofilms under flow conditions.

    abstract::Confocal microscopy-based monitoring of pH in biofilms is gaining increasing interest, as it allows for a quick assessment of horizontal pH gradients without mechanically perturbing the biofilm. Ratiometric monitoring of pH with the fluorescent dye C-SNARF-4 has been used to reliably map extracellular pH in the basal ...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2018.08.007

    authors: Schlafer S,Baelum V,Dige I

    更新日期:2018-09-01 00:00:00

  • Microscopic quantification of bacterial invasion by a novel antibody-independent staining method.

    abstract::Microscopic discrimination between extracellular and invasive, intracellular bacteria is a valuable technique in microbiology and immunology. We describe a novel fluorescence staining protocol, called FITC-biotin-avidin (FBA) staining, which allows the differentiation between extracellular and intracellular bacteria a...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2004.05.008

    authors: Agerer F,Waeckerle S,Hauck CR

    更新日期:2004-10-01 00:00:00

  • Use of Enterococcus faecalis and Bacillus atrophaeus as surrogates to establish and maintain laboratory proficiency for concentration of water samples using ultrafiltration.

    abstract::The U.S. Environmental Protection Agency's (EPA) Water Laboratory Alliance (WLA) currently uses ultrafiltration (UF) for concentration of biosafety level 3 (BSL-3) agents from large volumes (up to 100-L) of drinking water prior to analysis. Most UF procedures require comprehensive training and practice to achieve and ...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2015.08.013

    authors: Mapp L,Klonicki P,Takundwa P,Hill VR,Schneeberger C,Knee J,Raynor M,Hwang N,Chambers Y,Miller K,Pope M

    更新日期:2015-11-01 00:00:00

  • Developments in immunological technologies leading to improvements in point-of-care diagnostic testing.

    abstract::Rapid and accurate point of care testing is of great concern, especially in terms of detecting infectious disease outbreaks in developing countries having high population burdens. While numerous detection systems are currently available, care must be taken in choosing those that are reliable and have proven acceptable...

    journal_title:Journal of microbiological methods

    pub_type: 社论

    doi:10.1016/j.mimet.2011.05.015

    authors: Gurtler V,Pavia C

    更新日期:2011-09-01 00:00:00

  • Quantifying biofilm structure using image analysis.

    abstract::We have developed and implemented methods of extracting morphological features from images of biofilms in order to quantify the characteristics of the inherent heterogeneity. This is a first step towards quantifying the relationship between biofilm heterogeneity and the underlying processes, such as mass-transport dyn...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/s0167-7012(99)00097-4

    authors: Yang X,Beyenal H,Harkin G,Lewandowski Z

    更新日期:2000-01-01 00:00:00

  • Use of the frc gene as a molecular marker to characterize oxalate-oxidizing bacterial abundance and diversity structure in soil.

    abstract::Oxalate catabolism, which can have both medical and environmental implications, is performed by phylogenetically diverse bacteria. The formyl-CoA-transferase gene was chosen as a molecular marker of the oxalotrophic function. Degenerated primers were deduced from an alignment of frc gene sequences available in databas...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2008.09.020

    authors: Khammar N,Martin G,Ferro K,Job D,Aragno M,Verrecchia E

    更新日期:2009-02-01 00:00:00

  • Simplified amplified-fragment length polymorphism method for genotyping Mycobacterium tuberculosis isolates.

    abstract::A simplified amplified-fragment length polymorphism (AFLP) method was developed and applied to genotype 52 Mycobacterium tuberculosis isolates. This method can be carried out using only one restriction enzyme (XhoI), one double strand adapter, and one PCR primer. The amounts of DNA and DNA polymerase, and the concentr...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2009.07.007

    authors: Viader-Salvadó JM,Flores-Gracia J,Vega-Alonso AS,Treviño-Alvarado VM,Molina-Torres CA,Vera-Cabrera L,Guerrero-Olazarán M

    更新日期:2009-09-01 00:00:00

  • Use of an alternative Archaea-specific probe for methanogen detection.

    abstract::An alternative 16S rRNA-targeted oligonucleotide probe specific for Archaea was developed and used for detection of methanogens in anaerobic reactors. The designed probe was checked for its specificity by computer-aided comparative sequence analysis. For in situ application, optimal stringency conditions were adjusted...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2004.11.017

    authors: Jupraputtasri W,Boonapatcharoen N,Cheevadhanarak S,Chaiprasert P,Tanticharoen M,Techkarnjanaruk S

    更新日期:2005-04-01 00:00:00

  • Arsenic interception by cell wall of bacteria observed with surface-enhanced Raman scattering.

    abstract::The purpose of this study was to determine the interactions between arsenic (As) resistant bacteria and As, using surface-enhanced Raman scattering (SERS) and Fourier transform infrared (FTIR) spectroscopy. According to our 16S rDNA results, eight bacteria isolated from the environment can be identified to four genera...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2012.03.018

    authors: Tian H,Zhuang G,Ma A,Jing C

    更新日期:2012-06-01 00:00:00

  • Status of methods for assessing bacterial cell surface charge properties based on zeta potential measurements.

    abstract::Surface interfacial physiology is particularly important to unicellular organisms with regard to maintenance of optimal cell function. Bacterial cell surfaces possess net negative electrostatic charge by virtue of ionized phosphoryl and carboxylate substituents on outer cell envelope macromolecules which are exposed t...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章,评审

    doi:10.1016/s0167-7012(00)00224-4

    authors: Wilson WW,Wade MM,Holman SC,Champlin FR

    更新日期:2001-01-01 00:00:00

  • Development of a novel plasmid vector pTIO-1 adapted for electrotransformation of Porphyromonas gingivalis.

    abstract::We report here the construction of a plasmid vector designed for the efficient electrotransformation of the periodontal pathogen Porphyromonas gingivalis. The novel Escherichia coli-Bacteroides/P. gingivalis shuttle vector, designated pTIO-1, is based on the 11.0-kb E. coli-Bacteroides conjugative shuttle vector, pVAL...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2014.07.032

    authors: Tagawa J,Inoue T,Naito M,Sato K,Kuwahara T,Nakayama M,Nakayama K,Yamashiro T,Ohara N

    更新日期:2014-10-01 00:00:00

  • Rapid and simple detection of food poisoning bacteria by bead assay with a microfluidic chip-based system.

    abstract::A rapid bead assay for detecting pathogenic bacteria with a simple microfluidic chip-based system was developed. Five oligonucleotide probes corresponding to the 16S rRNA of the targeted bacteria were coupled covalently to fluorescent beads. Four species of bacteria (Escherichia coli, Salmonella enterica subsp. enteri...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2006.03.014

    authors: Ikeda M,Yamaguchi N,Tani K,Nasu M

    更新日期:2006-11-01 00:00:00

  • Methodological issues in the quantification of subgingival microorganisms using the checkerboard technique.

    abstract::The reproducibility and reliability of quantitative microbiological assessments using the DNA-DNA hybridization "checkerboard method" (CKB) were assessed. The data originated from 180 chronic periodontitis patients, who were enrolled in a clinical trial and sampled at baseline, and 3 and 12m post-therapy. The samples ...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2015.01.010

    authors: Dahlen G,Preus HR,Baelum V

    更新日期:2015-03-01 00:00:00

  • Pushing the limits for amplifying BrdU-labeled DNA encoding 16S rRNA: DNA polymerase as the determining factor.

    abstract::Identifying microorganisms that are active under specific conditions in ecosystems is a challenge in microbial ecology. Recently, the bromodeoxyuridine (BrdU) technique was developed to label actively growing cells. BrdU, a thymidine analog, is incorporated into newly synthesized DNA, and the BrdU-labeled DNA is then ...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2010.09.016

    authors: Roux-Michollet DD,Schimel JP,Holden PA

    更新日期:2010-12-01 00:00:00

  • Direct common gram-negative bacterial identification from positive blood culture bottles by SELDI-TOF MS.

    abstract::A protein database was constructed and validated with identification rate over 90% for the 4 most common Gram-negative bacteria on agar plates. By protein masses comparison, 120 bacteria of the 4 species from blood culture bottles were identified. The concordance was high (Kappa=0.906) between our method and conventio...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2014.07.021

    authors: Xiao D,Yang Y,Jiang W,Zhang H,Liu H,Yu H,Xie C,Zhong M,Chen L,Huang W

    更新日期:2014-10-01 00:00:00

  • Development of RNA aptamers for detection of Salmonella Enteritidis.

    abstract::We developed and evaluated RNA aptamers to analyze their potential for use in detecting Salmonella Enteritidis. The selected aptamer was observed to specifically bind to Salmonella Enteritidis without any cross-reactivity to other Salmonella serovars. Thus, this study suggests that aptamers specific to Salmonella Ente...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2012.01.014

    authors: Hyeon JY,Chon JW,Choi IS,Park C,Kim DE,Seo KH

    更新日期:2012-04-01 00:00:00

  • RNA-based sandwich hybridisation method for detection of lactic acid bacteria in brewery samples.

    abstract::Recently we showed the applicability and sensitivity of the RNA-based sandwich hybridisation assay (SHA) for detection of gram-negative cells in environmental samples [Leskelä, T., Tilsala-Timisjärvi, A., Kusnetsov, J., Neubauer, P., Breitenstein, A., 2005. Sensitive genus-specific detection of Legionella by a 16S rRN...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2006.10.009

    authors: Huhtamella S,Leinonen M,Nieminen T,Fahnert B,Myllykoski L,Breitenstein A,Neubauer P

    更新日期:2007-03-01 00:00:00

  • Development of an oriC vector for use in Mycoplasma synoviae.

    abstract::Mycoplasma synoviae, an important poultry pathogen, belonging to the class Mollicutes, causes airsacculitis, synovitis, decreased egg production and produces significant economic losses. Efforts to determine M. synoviae virulence factors and their role in pathogenicity require suitable tools for genetic manipulation o...

    journal_title:Journal of microbiological methods

    pub_type: 杂志文章

    doi:10.1016/j.mimet.2014.05.014

    authors: Shahid MA,Marenda MS,Markham PF,Noormohammadi AH

    更新日期:2014-08-01 00:00:00