A simple method for on-gel detection of myrosinase activity

Sándor Gonda, Zsolt Sz ucs, Tamás Plaszkó, Zoltán Cziáky, Attila Kiss-Szikszai, G. Vasas, Márta M-Hamvas

Research output: Contribution to journalArticle

2 Citations (Scopus)

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.

Original languageEnglish
Article number2204
JournalMolecules
Volume23
Issue number9
DOIs
Publication statusPublished - Aug 31 2018

Fingerprint

Gels
vegetables
gels
Glucosinolates
Sulfatases
Vegetables
Armoracia
enzymes
proteins
fungi
nitriles
Sinapis
activity (biology)
Isothiocyanates
food
purification
Native Polyacrylamide Gel Electrophoresis
Spices
reagents
Nitriles

Keywords

  • Desulfo-sinigrin
  • LC-ESI-MS
  • Myrosinase
  • On-gel detection
  • Sulfatase
  • Thioglucosidase

ASJC Scopus subject areas

  • Analytical Chemistry
  • Chemistry (miscellaneous)
  • Molecular Medicine
  • Pharmaceutical Science
  • Drug Discovery
  • Physical and Theoretical Chemistry
  • Organic Chemistry

Cite this

Gonda, S., Sz ucs, Z., Plaszkó, T., Cziáky, Z., Kiss-Szikszai, A., Vasas, G., & M-Hamvas, M. (2018). A simple method for on-gel detection of myrosinase activity. Molecules, 23(9), [2204]. https://doi.org/10.3390/molecules23092204

A simple method for on-gel detection of myrosinase activity. / Gonda, Sándor; Sz ucs, Zsolt; Plaszkó, Tamás; Cziáky, Zoltán; Kiss-Szikszai, Attila; Vasas, G.; M-Hamvas, Márta.

In: Molecules, Vol. 23, No. 9, 2204, 31.08.2018.

Research output: Contribution to journalArticle

Gonda, S, Sz ucs, Z, Plaszkó, T, Cziáky, Z, Kiss-Szikszai, A, Vasas, G & M-Hamvas, M 2018, 'A simple method for on-gel detection of myrosinase activity', Molecules, vol. 23, no. 9, 2204. https://doi.org/10.3390/molecules23092204
Gonda S, Sz ucs Z, Plaszkó T, Cziáky Z, Kiss-Szikszai A, Vasas G et al. A simple method for on-gel detection of myrosinase activity. Molecules. 2018 Aug 31;23(9). 2204. https://doi.org/10.3390/molecules23092204
Gonda, Sándor ; Sz ucs, Zsolt ; Plaszkó, Tamás ; Cziáky, Zoltán ; Kiss-Szikszai, Attila ; Vasas, G. ; M-Hamvas, Márta. / A simple method for on-gel detection of myrosinase activity. In: Molecules. 2018 ; Vol. 23, No. 9.
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