Next Article in Journal
Application of Dendrimers for the Treatment of Infectious Diseases
Previous Article in Journal
Investigation of Novel Pesticides with Insecticidal and Antifungal Activities: Design, Synthesis and SAR Studies of Benzoylpyrimidinylurea Derivatives
Previous Article in Special Issue
First Look at the Venom of Naja ashei
Article Menu
Issue 9 (September) cover image

Export Article

Open AccessArticle
Molecules 2018, 23(9), 2204; https://doi.org/10.3390/molecules23092204

A Simple Method for On-Gel Detection of Myrosinase Activity

1
Department of Botany, Division of Pharmacognosy, University of Debrecen, Egyetem tér 1, H-4010 Debrecen, Hungary
2
Agricultural and Molecular Research and Service Institute, University of Nyíregyháza, Sóstói str. 31/b, H-4400 Nyíregyháza, Hungary
3
Department of Organic Chemistry, University of Debrecen, Egyetem tér 1, H-4010 Debrecen, Hungary
*
Author to whom correspondence should be addressed.
Received: 5 July 2018 / Revised: 27 August 2018 / Accepted: 28 August 2018 / Published: 31 August 2018
(This article belongs to the Special Issue Analysis of Peptides and Proteins by Electrophoretic Techniques)
Full-Text   |   PDF [1271 KB, uploaded 31 August 2018]   |  

Abstract

Myrosinase is an enzyme present in many functional foods and spices, particularly in Cruciferous vegetables. It hydrolyses glucosinolates which thereafter rearrange into bioactive volatile constituents (isothiocyanates, nitriles). We aimed to develop a simple reversible method for on-gel detection of myrosinase. Reagent composition and application parameters for native PAGE and SDS-PAGE gels were optimized. The proposed method was successfully applied to detect myrosinase (or sulfatase) on-gel: the detection solution contains methyl red which gives intensive red bands where the HSO4 is enzymatically released from the glucosinolates. Subsequently, myrosinase was successfully distinguished from sulfatase by incubating gel bands in a derivatization solution and examination by LC-ESI-MS: myrosinase produced allyl isothiocyanate (detected in conjugate form) while desulfo-sinigrin was released by sulfatase, as expected. After separation of 80 µg protein of crude extracts of Cruciferous vegetables, intensive color develops within 10 min. On-gel detection was found to be linear between 0.031–0.25 U (pure Sinapis alba myrosinase, R2 = 0.997). The method was successfully applied to detection of myrosinase isoenzymes from horseradish, Cruciferous vegetables and endophytic fungi of horseradish as well. The method was shown to be very simple, rapid and efficient. It enables detection and partial characterization of glucosinolate decomposing enzymes without protein purification. View Full-Text
Keywords: myrosinase; thioglucosidase; sulfatase; on-gel detection; desulfo-sinigrin; LC-ESI-MS myrosinase; thioglucosidase; sulfatase; on-gel detection; desulfo-sinigrin; LC-ESI-MS
Figures

Figure 1

This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

Supplementary material

SciFeed

Share & Cite This Article

MDPI and ACS Style

Gonda, S.; Szűcs, Z.; Plaszkó, T.; Cziáky, Z.; Kiss-Szikszai, A.; Vasas, G.; M-Hamvas, M. A Simple Method for On-Gel Detection of Myrosinase Activity. Molecules 2018, 23, 2204.

Show more citation formats Show less citations formats

Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Related Articles

Article Metrics

Article Access Statistics

1

Comments

[Return to top]
Molecules EISSN 1420-3049 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert
Back to Top