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Purification and kinetic characterization of an anionic peroxidase from melon (Cucumis melo L.) cultivated under
J N Rodríguez-López1, J C Espín, F del Amor
1Grupo de Investigación de Enzimología (GENZ), Departamento de Bioquímica y Biología Molecular A, Facultad de Biología, Universidad de Murcia, E-30100 Espinardo, Murcia, Spain.
Abstract:
The partial characterization of an anionic peroxidase in melon fruit is described. Four melon peroxidase (MPX) isoenzymes were detected in crude extracts after isoelectric focusing. The major MPX isoenzyme (pI = 3.7) was partially purified by including hydrophobic and anion-exchange chromatography in the purification scheme. The sample obtained was used to characterize MPX. This peroxidase did not show activity on ascorbic acid but oxidized guaiacol at a high rate, showing an optimum pH of 5.5 when acting on this last reducing substrate. Melon fruits grown under highly saline conditions showed slightly increased levels of this anionic isoenzyme. Kinetic studies using 2,2'-azinobis(3-ethylbenzothiazolinesulfonic acid) (ABTS) as reducing substrate showed that increased salinity in the growth medium did not modify the kinetic parameters of melon peroxidase on both hydrogen peroxide and reducing substrate.
Insights
Researchers partially characterized melon fruit anionic peroxidase (MPX). Salinity increased MPX levels but did not alter its enzymatic activity kinetics, suggesting robust performance in challenging environments.
Area of Science:
- Plant Biochemistry
- Enzymology
- Fruit Science
Background:
- Peroxidases play crucial roles in plant defense and fruit ripening.
- Understanding melon fruit peroxidase (MPX) is vital for agricultural applications.
- Environmental stresses like salinity can impact fruit enzyme activity.
Purpose of the Study:
- To partially characterize an anionic peroxidase in melon fruit.
- To investigate the effect of salinity on MPX levels and kinetics.
Main Methods:
- Isoelectric focusing to detect MPX isoenzymes.
- Hydrophobic and anion-exchange chromatography for partial purification.
- Enzyme activity assays using guaiacol and ABTS as substrates.
- Kinetic parameter determination under varying salinity conditions.
Main Results:
- Four MPX isoenzymes were identified, with the major one having a pI of 3.7.
- The partially purified MPX efficiently oxidized guaiacol at an optimal pH of 5.5.
- Melon fruits under saline conditions exhibited slightly elevated levels of the anionic MPX isoenzyme.
- Salinity did not significantly alter the kinetic parameters of MPX for hydrogen peroxide or ABTS reduction.
Conclusions:
- Melon fruit possesses a distinct anionic peroxidase with specific substrate preferences.
- MPX shows a degree of resilience to salinity stress, with increased expression but unchanged catalytic efficiency.
- These findings contribute to understanding plant enzyme responses to environmental factors.