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  1. Article ; Online: Recombinant laccase from Pediococcus acidilactici CECT 5930 with ability to degrade tyramine.

    Callejón, Sara / Sendra, Ramón / Ferrer, Sergi / Pardo, Isabel

    PloS one

    2017  Volume 12, Issue 10, Page(s) e0186019

    Abstract: Biogenic amines degradation by bacterial laccases is little known, so we have cloned and heterologously expressed, in E. coli, a new laccase from Pediococcus acidilactici CECT 5930 (Lpa5930), a lactic acid bacterium commonly found in foods able to ... ...

    Abstract Biogenic amines degradation by bacterial laccases is little known, so we have cloned and heterologously expressed, in E. coli, a new laccase from Pediococcus acidilactici CECT 5930 (Lpa5930), a lactic acid bacterium commonly found in foods able to degrade tyramine. The recombinant enzyme has been characterized by physical and biochemical assays. Here we report the optimization of expression and purification procedures of this laccase. DNA encoding sequence of laccase from P. acidilactici was amplified by PCR and cloned into the expression plasmid pET28a for induction by isopropyl-β-D-thiogalactoipyranoside. Protein expression was performed in E. coli BL21(DE3) harboring pGro7 plasmid expressing a chaperone folding assistant induced by arabinose. Purification was performed by column metal-chelating chromatography on Ni-NTA-agarose. The laccase enzyme obtained has an apparent molecular mass of ∼60 kDa, an optimum temperature activity toward 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) of 28°C, and was quickly inactivated at temperatures higher than 70°C. The apparent Km value for ABTS was 1.7 mM and the Vmax obtained was 24 U/mg. In addition to ABTS, recombinant Lpa5930 laccase degraded the biogenic amine tyramine at pH 9.5 and pH 4.0 with or without ABTS as a mediator. Tyramine degradation by laccases could solve the problems generated in food due to the presence of this toxic compound.
    MeSH term(s) Amino Acid Sequence ; Bacterial Proteins/chemistry ; Base Sequence ; Benzothiazoles/metabolism ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Hydrogen-Ion Concentration ; Laccase/metabolism ; Oxidation-Reduction ; Pediococcus acidilactici/enzymology ; Recombinant Proteins/isolation & purification ; Recombinant Proteins/metabolism ; Sequence Analysis, DNA ; Spectrophotometry, Ultraviolet ; Substrate Specificity ; Sulfonic Acids/metabolism ; Temperature ; Tyramine/metabolism
    Chemical Substances Bacterial Proteins ; Benzothiazoles ; Recombinant Proteins ; Sulfonic Acids ; 2,2'-azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid (28752-68-3) ; Laccase (EC 1.10.3.2) ; Tyramine (X8ZC7V0OX3)
    Language English
    Publishing date 2017-10-11
    Publishing country United States
    Document type Journal Article
    ZDB-ID 2267670-3
    ISSN 1932-6203 ; 1932-6203
    ISSN (online) 1932-6203
    ISSN 1932-6203
    DOI 10.1371/journal.pone.0186019
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: Structural analysis and biochemical properties of laccase enzymes from two Pediococcus species.

    Olmeda, Isidoro / Casino, Patricia / Collins, Robert E / Sendra, Ramón / Callejón, Sara / Huesa, Juanjo / Soares, Alexei S / Ferrer, Sergi / Pardo, Isabel

    Microbial biotechnology

    2021  Volume 14, Issue 3, Page(s) 1026–1043

    Abstract: Prokaryotic laccases are emergent biocatalysts. However, they have not been broadly found and characterized in bacterial organisms, especially in lactic acid bacteria. Recently, a prokaryotic laccase from the lactic acid bacterium Pediococcus ... ...

    Abstract Prokaryotic laccases are emergent biocatalysts. However, they have not been broadly found and characterized in bacterial organisms, especially in lactic acid bacteria. Recently, a prokaryotic laccase from the lactic acid bacterium Pediococcus acidilactici 5930, which can degrade biogenic amines, was discovered. Thus, our study aimed to shed light on laccases from lactic acid bacteria focusing on two Pediococcus laccases, P. acidilactici 5930 and Pediococcus pentosaceus 4816, which have provided valuable information on their biochemical activities on redox mediators and biogenic amines. Both laccases are able to oxidize canonical substrates as ABTS, ferrocyanide and 2,6-DMP, and non-conventional substrates as biogenic amines. With ABTS as a substrate, they prefer an acidic environment and show sigmoidal kinetic activity, and are rather thermostable. Moreover, this study has provided the first structural view of two lactic acid bacteria laccases, revealing new structural features not seen before in other well-studied laccases, but which seem characteristic for this group of bacteria. We believe that understanding the role of laccases in lactic acid bacteria will have an impact on their biotechnological applications and provide a framework for the development of engineered lactic acid bacteria with enhanced properties.
    MeSH term(s) Bacteria/metabolism ; Laccase/metabolism ; Oxidation-Reduction ; Pediococcus/metabolism ; Prokaryotic Cells/metabolism
    Chemical Substances Laccase (EC 1.10.3.2)
    Language English
    Publishing date 2021-02-26
    Publishing country United States
    Document type Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S.
    ZDB-ID 2406063-X
    ISSN 1751-7915 ; 1751-7915
    ISSN (online) 1751-7915
    ISSN 1751-7915
    DOI 10.1111/1751-7915.13751
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  3. Article: Ability of Kocuria varians LTH 1540 To Degrade Putrescine: Identification and Characterization of a Novel Amine Oxidase

    Callejon, Sara / Ramon Sendra / Sergi Ferrer / Isabel Pardo

    Journal of agricultural and food chemistry. 2015 Apr. 29, v. 63, no. 16

    2015  

    Abstract: This work describes the identification and characterization of an amine oxidase from Kocuria varians LTH 1540 (syn. Micrococcus varians) primarily acting on putrescine. Data from MALDI-TOF MS/MS and the identification of Δ1-pyrroline as degradation ... ...

    Abstract This work describes the identification and characterization of an amine oxidase from Kocuria varians LTH 1540 (syn. Micrococcus varians) primarily acting on putrescine. Data from MALDI-TOF MS/MS and the identification of Δ1-pyrroline as degradation product from putrescine indicate that the enzyme is a flavin-dependent putrescine oxidase (PuO). Properties of partially purified enzyme have been determined. The enzyme oxidizes diamines, putrescine and cadaverine, and, to a lesser extent, polyamines, such as spermidine, but not monoamines. The kinetic constants (Km and Vmax) for the two major substrates were 94 ± 10 μM and 2.3 ± 0.1 μmol/min·mg for putrescine and 75 ± 5 μM and 0.15 ± 0.02 μmol/min·mg for cadaverine. Optimal temperature and pH were 45 °C and 8.5, respectively. Enzyme was stable until 50 °C. K. varians PuO is sensitive to human flavin-dependent amine oxidase inhibitors and carboxyl-modifying compounds. The new enzyme has been isolated from a bacterial starter used in the manufacture of fermented meat. One of the problems of fermented foods or beverages is the presence of toxic biogenic amines produced by bacteria. The importance of this works lies in the description of a new enzyme able to degrade two of the most abundant biogenic amines (putrescine and cadaverine), the use of which could be envisaged to diminish biogenic amines content in foods in the future.
    Keywords Kocuria varians ; bacteria ; beverages ; cadaverine ; enzyme stability ; fermented meat ; humans ; manufacturing ; matrix-assisted laser desorption-ionization mass spectrometry ; monoamines ; pH ; putrescine ; spermidine ; temperature ; toxicity
    Language English
    Dates of publication 2015-0429
    Size p. 4170-4178.
    Publishing place American Chemical Society, Books and Journals Division
    Document type Article
    ZDB-ID 241619-0
    ISSN 1520-5118 ; 0021-8561
    ISSN (online) 1520-5118
    ISSN 0021-8561
    DOI 10.1021/jf5026967
    Database NAL-Catalogue (AGRICOLA)

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    Z 1185: Show issues

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  4. Article ; Online: Ability of Kocuria varians LTH 1540 To Degrade Putrescine: Identification and Characterization of a Novel Amine Oxidase.

    Callejón, Sara / Sendra, Ramón / Ferrer, Sergi / Pardo, Isabel

    Journal of agricultural and food chemistry

    2015  Volume 63, Issue 16, Page(s) 4170–4178

    Abstract: This work describes the identification and characterization of an amine oxidase from Kocuria varians LTH 1540 (syn. Micrococcus varians) primarily acting on putrescine. Data from MALDI-TOF MS/MS and the identification of Δ(1)-pyrroline as degradation ... ...

    Abstract This work describes the identification and characterization of an amine oxidase from Kocuria varians LTH 1540 (syn. Micrococcus varians) primarily acting on putrescine. Data from MALDI-TOF MS/MS and the identification of Δ(1)-pyrroline as degradation product from putrescine indicate that the enzyme is a flavin-dependent putrescine oxidase (PuO). Properties of partially purified enzyme have been determined. The enzyme oxidizes diamines, putrescine and cadaverine, and, to a lesser extent, polyamines, such as spermidine, but not monoamines. The kinetic constants (Km and Vmax) for the two major substrates were 94 ± 10 μM and 2.3 ± 0.1 μmol/min·mg for putrescine and 75 ± 5 μM and 0.15 ± 0.02 μmol/min·mg for cadaverine. Optimal temperature and pH were 45 °C and 8.5, respectively. Enzyme was stable until 50 °C. K. varians PuO is sensitive to human flavin-dependent amine oxidase inhibitors and carboxyl-modifying compounds. The new enzyme has been isolated from a bacterial starter used in the manufacture of fermented meat. One of the problems of fermented foods or beverages is the presence of toxic biogenic amines produced by bacteria. The importance of this works lies in the description of a new enzyme able to degrade two of the most abundant biogenic amines (putrescine and cadaverine), the use of which could be envisaged to diminish biogenic amines content in foods in the future.
    MeSH term(s) Bacterial Proteins/chemistry ; Bacterial Proteins/genetics ; Bacterial Proteins/metabolism ; Biodegradation, Environmental ; Enzyme Stability ; Hydrogen-Ion Concentration ; Kinetics ; Micrococcus/chemistry ; Micrococcus/enzymology ; Micrococcus/genetics ; Oxidoreductases Acting on CH-NH Group Donors/chemistry ; Oxidoreductases Acting on CH-NH Group Donors/genetics ; Oxidoreductases Acting on CH-NH Group Donors/metabolism ; Putrescine/metabolism
    Chemical Substances Bacterial Proteins ; putrescine oxidase (EC 1.4.3.10) ; Oxidoreductases Acting on CH-NH Group Donors (EC 1.5.-) ; Putrescine (V10TVZ52E4)
    Language English
    Publishing date 2015-04-29
    Publishing country United States
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 241619-0
    ISSN 1520-5118 ; 0021-8561
    ISSN (online) 1520-5118
    ISSN 0021-8561
    DOI 10.1021/jf5026967
    Database MEDical Literature Analysis and Retrieval System OnLINE

    Full text online

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    In stock of ZB MED Cologne/Königswinter

    Zs.A 1172: Show issues Location:
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