3-Methyl-2-benzothiazolinone hydrazone and 3-dimethylamino benzoic acid as substrates for the development of polyphenoloxidase and phenoloxidase activity by zymograms
| Autor: | A I Marina-Ramírez, Jorge Álvarez-Cervantes, E Aguirre-von Wobeser, Miguel Angel Anducho-Reyes, Yuridia Mercado-Flores, Y. García-Esquivel, Alejandro Téllez-Jurado |
|---|---|
| Přispěvatelé: | Consejo Nacional de Ciencia y Tecnología (México) |
| Rok vydání: | 2020 |
| Předmět: |
Laccase
DMAB Chromatography ABTS biology Chemistry MBTH Zymography Lentinus strigosus Lignin peroxidase Environmental Science (miscellaneous) biology.organism_classification Agricultural and Biological Sciences (miscellaneous) Staining Isoenzymes chemistry.chemical_compound Phenoloxidase Original Article Guaiacol Biotechnology Benzoic acid |
| Zdroj: | 3 Biotech Digital.CSIC. Repositorio Institucional del CSIC instname |
| ISSN: | 2190-572X |
| Popis: | In the present study, a sequential staining process of polyphenoloxidase and phenoloxidase enzymes was designed by the zymography technique. As a first step, electrophoresis was carried out under native conditions, and later, first staining was carried out with a revealing solution of 3-methyl-2-benzothiazoline hydrazone (MBTH)—3-dimethylamino benzoic acid (DMAB) that allowed the visualization of polyphenoloxidase enzymes, and later and using the same gel, we proceeded to the differential staining of phenoloxidase, adding a solution of HO. The technique was standardized using commercial enzymes of laccase (T. versicolor) and horseradish. The technique was used to identify polyphenoloxidases (laccases) and phenoloxidases (lignin peroxidase) of crude extracts obtained from the growth of the basidiomycete Lentinus strigosus on Pinus radiata. The technique showed great sensitivity to detect the different enzymatic activities (1.56 Activity Unit/mL minimum) in the same gel without interference between the enzymes and the solutions used. On the other hand, the efficiency of the technique was compared with the substrates that are commonly used for the detection of this type of activities such as 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) and guaiacol, observing greater sensitivity and minimal interference, so that the present method will allow in the same gel, and visualize polyphenoloxidase and phenoloxidase activities simultaneously facilitating expression studies. National Council of Science and Technology (CONACyT) of Mexico |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full text from SpringerLink