O-GlcNAcase catalyzes the removal of N-acetylglucosamine residues from serine and threonine residues of post-translationally modified proteins using a catalytic mechanism involving substrate-assisted catalysis and general acid/base catalysis. Since thioglycosides are widely perceived as resistant to hydrolysis by glycosidases, it was surprising to find that O-GlcNAcase also catalyzes the efficient hydrolysis of S-glycosides. Bronsted analyses and pH-activity studies of the O-GlcNAcase-catalyzed hydrolysis of a series of aryl S- and O-glycosides reveal that O-GlcNAcase effects hydrolysis of thioglycosides without the assistance of general acid catalysis. α-Deuterium kinetic isotope effects for O- and S-glycosides, as well as Taft-like analyses using N-fluoroacetyl-β-glycosides, suggest that O-GlcNAcase accomplishes hydrolysis of thioglycosides by stabilizing late transition states. For S-glycosides this transition state shows greater nucleophilic participation from the 2-acetamido group than for O-glycosid...

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O-GlcNAcase catalyzes cleavage of thioglycosides without general acid catalysis.

Most known biological roles of the glucosinolate–myrosinase system depend on its functioning as a ‘chemical bomb’. This term describes chemical defences which are based on the accumulation of inert components in separate compartments and rapid release of large amounts of defensive chemicals through mixing of these components and their reaction with each other upon tissue disruption. The essential components of the ‘chemical bomb’ formed by the glucosinolate–myrosinase system are the water-soluble glucosinolates and their hydrolytic enzymes, myrosinases. The defensive chemicals released upon mixing of glucosinolates and myrosinases are the rather lipophilic, irritant and toxic isothiocyanates (mustard oils; reflected in the term ‘mustard oil bomb’) and their derivatives. In addition, so-called specifier proteins may promote the formation of alternative products such as nitriles and epithionitriles. As a consequence, tissue disruption typically leads to the release of complex mixtures of glucosinolate breakdown products. Apart from directly defensive roles (e.g., as toxins), the breakdown products may also be involved in indirect defences (e.g., as signals in tritrophic interactions). Besides glucosinolate breakdown upon tissue disruption, their turnover in intact tissue has been discussed to contribute sulphur and nitrogen for plant growth. In addition, breakdown of glucosinolates in nondisrupted tissue has been observed upon pathogen attack. This chapter summarizes the current knowledge on the protein components of the glucosinolate breakdown machinery and its compartmentation, highlights the relevance of different breakdown product types and their regulated formation, and discusses pathways and roles of glucosinolate breakdown in nondisrupted tissue.

Chapter Six – Glucosinolate Breakdown

Glucosinolate-myrosinase is a substrate-enzyme defense mechanism present in Brassica crops. This binary system provides the plant with an efficient system against herbivores and pathogens. For humans, it is well known for its anti-carcinogenic, anti-inflammatory, immunomodulatory, anti-bacterial, cardio-protective, and central nervous system protective activities. Glucosinolate and myrosinase are spatially present in different cells that upon tissue disruption come together and result in the formation of a variety of hydrolysis products with diverse physicochemical and biological properties. The myrosinase-catalyzed reaction starts with cleavage of the thioglucosidic linkage resulting in release of a D-glucose and an unstable thiohydroximate-O-sulfate. The outcome of this thiohydroximate-O-sulfate has been shown to depend on the structure of the glucosinolate side chain, the presence of supplementary proteins known as specifier proteins and/or on the physiochemical condition. Myrosinase was first reported in mustard seed during 1939 as a protein responsible for release of essential oil. Until this date, myrosinases have been characterized from more than 20 species of Brassica, cabbage aphid, and many bacteria residing in the human intestine. All the plant myrosinases are reported to be activated by ascorbic acid while aphid and bacterial myrosinases are found to be either neutral or inhibited. Myrosinase catalyzes hydrolysis of the S-glycosyl bond, O-β glycosyl bond, and O-glycosyl bond. This review summarizes information on myrosinase, an essential component of this binary system, including its structural and molecular properties, mechanism of action, and its regulation and will be beneficial for the research going on the understanding and betterment of the glucosinolate-myrosinase system from an ecological and nutraceutical perspective.

Myrosinase: insights on structural, catalytic, regulatory, and environmental interactions.

Beta-glucosidase inhibitors are being extensively studied for use as anti-diabetics, anti-obesity and anti-tumour compounds. So far, these compounds have been reported in large numbers from plants, mushrooms, algae and fungi. There are very few reports of such inhibitors from bacteria in the open literature, particularly marine bacteria; although the best known inhibitor deoxynojirimycin was isolated from bacilli and actinomycete. Through this study, we tried to discover the diversity of microbial associates of marine sponge and sediment producing β-glucosidase inhibitors. We found 41 (22.7%) out of 181 bacteria, produced such inhibitors. The inhibitors are abundant in bacterial associates of marine sponge Aka coralliphaga. When these bacteria were phylogenetically analyzed, it was found that marine bacteria producing glucosidase inhibitors belong to the phylum Firmicutes (23), Actinobacteria (9), Proteobacteria (7) and Bacteroidetes (1). A significant number of marine bacteria belonging to a wide range of bacterial taxa were found to produce β-glucosidase inhibitors. These compounds are abundantly present in bacteria of the phylum Firmicutes followed by the phylum Actinobacteria. The results nurture a hope of finding new compounds, which can inhibit glucosidases, in the bacterial domain of marine organisms. Thus, marine microbial cells can be utilized as producers of pharmacologically essential enzyme inhibitors.

https://link.springer.com/content/pdf/10.1186%2F1475-2859-12-35.pdf

Diversity of marine bacteria producing beta-glucosidase inhibitors.

Glucosinolate preservation in stored Brassicaceae seed meals

Screening for novel thioglucoside hydrolase activity resulted in the isolation of Sphingobacterium sp. strain OTG1 from enrichment cultures containing octylthioglucoside (OTG). OTG was hydrolysed into octanethiol and glucose by cell free extracts. Besides thioglucoside hydrolysis, several other glucoside hydrolase activities were detected in the Sphingobacterium sp. strain OTG1 cell free extract. By adding β-glucosidase inhibitors it was possible to discriminate between these different activities. Ascorbic acid and D-gluconic acid lactone inhibited the hydrolysis of p-nitrophenyl β-glucoside, but did not affect octyl- and octylthioglucoside hydrolase activity. Besides OTG, various other thioglucosides were hydrolysed by the novel thioglucosidase, with almost the same activities regardless of the nature of the aglycone, including the myrosinase model substrate sinigrin (a glucosinolate). Sinigrin could also be used as a growth substrate by Sphingobacterium sp. strain OTG1, although at concentrations exceeding 0.15 mM degradation was not complete.

S. Hartmans

Papers

Thioglucosidase activity from Sphingobacterium sp. strain OTG1