An in situ activity assay for lysyl oxidases.

Amino Acid Oxidoreductases / genetics Animals Aorta / enzymology Biocatalysis Blotting, Western Cell Line Enzyme Assays / methods Fluorometry / methods Humans Male Mice, Inbred C57BL Mice, Knockout Protein-Lysine 6-Oxidase / genetics Rats

Journal

Communications biology
ISSN: 2399-3642
Titre abrégé: Commun Biol
Pays: England
ID NLM: 101719179

Informations de publication

Date de publication:
05 07 2021
Historique:
received: 05 04 2020
accepted: 16 06 2021
entrez: 6 7 2021
pubmed: 7 7 2021
medline: 17 8 2021
Statut: epublish

Résumé

The lysyl oxidase family of enzymes (LOXs) catalyze oxidative deamination of lysine side chains on collagen and elastin to initialize cross-linking that is essential for the formation of the extracellular matrix (ECM). Elevated expression of LOXs is highly associated with diverse disease processes. To date, the inability to detect total LOX catalytic function in situ has limited the ability to fully elucidate the role of LOXs in pathobiological mechanisms. Using LOXL2 as a representative member of the LOX family, we developed an in situ activity assay by utilizing the strong reaction between hydrazide and aldehyde to label the LOX-catalyzed allysine (-CHO) residues with biotin-hydrazide. The biotinylated ECM proteins are then labeled via biotin-streptavidin interaction and detected by fluorescence microscopy. This assay detects the total LOX activity in situ for both overexpressed and endogenous LOXs in cells and tissue samples and can be used for studies of LOXs as therapeutic targets.

Identifiants

DOI: 10.1038/s42003-021-02354-0 PMID: 34226627 PMC: PMC8257687
pubmed: 34226627
doi: 10.1038/s42003-021-02354-0
pii: 10.1038/s42003-021-02354-0
pmc: PMC8257687
doi:

Substances chimiques

Amino Acid Oxidoreductases EC 1.4.-
Loxl2 protein, mouse EC 1.4.3.-
Protein-Lysine 6-Oxidase EC 1.4.3.13

Types de publication

Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Langues

eng

Sous-ensembles de citation

IM

Pagination

840

Subventions

Organisme : NHLBI NIH HHS
ID : K08 HL145132
Pays : United States
Organisme : NHLBI NIH HHS
ID : R01 HL148112
Pays : United States

Commentaires et corrections

Type : ErratumIn

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Auteurs

Huilei Wang (H)

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA.

Alan Poe (A)

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA.

Lydia Pak (L)

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA.

Kavitha Nandakumar (K)

Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University, Baltimore, MD, USA.

Sandeep Jandu (S)

Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University, Baltimore, MD, USA.

Jochen Steppan (J)

Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University, Baltimore, MD, USA.
ORCID: 0000-0001-9856-512X

Reik Löser (R)

Institute of Radiopharmaceutical Cancer Research, Helmholtz-Zentrum Dresden-Rossendorf, Dresden, Germany.
ORCID: 0000-0003-1531-7601

Lakshmi Santhanam (L)

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA. LSantha1@jh.edu.
Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University, Baltimore, MD, USA. LSantha1@jh.edu.
Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, MD, USA. LSantha1@jh.edu.
ORCID: 0000-0002-2097-9397

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Classifications MeSH

questionsmedicales.fr › Enzymes et coenzymes › Enzymes › Oxidoreductases › Oxidoreductases acting on CH-NH2 group donors › Amino-acid oxidoreductases › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Eucaryotes › Animaux › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Système cardiovasculaire › Vaisseaux sanguins › Artères › Aorte › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Métabolisme › Biocatalyse › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Techniques d'investigation › Techniques de sonde moléculaire › Dosage immunologique › Immunotransfert › Technique de Western › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Cellules › Cellules cultivées › Lignée cellulaire › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Techniques d'investigation › Techniques de chimie analytique › Dosages enzymatiques › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Techniques d'investigation › Techniques de chimie analytique › Photométrie › Mesures de luminescence › Fluorimétrie › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Eucaryotes › Animaux › Chordés › Vertébrés › Mammifères › Eutheria › Primates › Haplorhini › Catarrhini › Hominidae › Humains › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Eucaryotes › Animaux › Chordés › Vertébrés › Mammifères › Eutheria › Rodentia › Muridae › Murinae › Souris › Lignées consanguines de souris › Souris de lignée C57BL › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Eucaryotes › Animaux › Chordés › Vertébrés › Mammifères › Eutheria › Rodentia › Muridae › Murinae › Souris › Souris transgéniques › Souris knockout › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Enzymes et coenzymes › Enzymes › Oxidoreductases › Oxidoreductases acting on CH-NH2 group donors › Amino-acid oxidoreductases › Lysyloxidase › An in situ activity assay for lysyl oxidases.
questionsmedicales.fr › Eucaryotes › Animaux › Chordés › Vertébrés › Mammifères › Eutheria › Rodentia › Muridae › Murinae › Rats › An in situ activity assay for lysyl oxidases.