Identification and characterization of a novel β-D-galactosidase that releases pyruvylated galactose

Higuchi, Yujiro; Matsufuji, Hitomi; Tanuma, Masanari; Arakawa, T.; Mori, Kazuki; Yamada, Chihaya; Shofia, Risa; Matsunaga, Eiji; Tashiro, Kosuke; Fushinobu, Shinya; Takegawa, Kaoru

doi:10.1038/s41598-018-30508-4

Cited by 10 publications

(5 citation statements)

References 31 publications

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“…Tyr 241 interacts hydrophobically with GlcA. The catalytic sites of carbohydrate hydrolases contain aromatic amino acid residues that fix the sugar ring via hydrophobic stacking interactions [30–33]. The aromatic amino acid residues participating in the hydrophobic stacking are often conserved within the same GH family (Fig.…”

Section: Resultsmentioning

confidence: 99%

Biochemical and structural characterization of a novel 4‐O‐α‐l‐rhamnosyl‐β‐d‐glucuronidase from Fusarium oxysporum

et al. 2021

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In this study, we have isolated the novel enzyme 4‐O‐α‐l‐rhamnosyl‐β‐d‐glucuronidase (FoBGlcA), which releases α‐l‐rhamnosyl (1→4) glucuronic acid from gum arabic (GA), from Fusarium oxysporum 12S culture supernatant, and for the first time report an enzyme with such catalytic activity. The gene encoding FoBGlcA was cloned and expressed in Pichia pastoris. When GA was subjected to the recombinant enzyme, > 95% of the l‐rhamnose (Rha) and d‐glucuronic acid in the substrate were released, which indicates that almost all Rha binds to the glucuronic acid at the end of the GA side chains. The crystal structure of FoBGlcA was determined using a single‐wavelength anomalous dispersion at 1.51 Å resolution. FoBGlcA consisted of an N‐terminal (β/α)8‐barrel domain and a C‐terminal antiparallel β‐sheet domain. This configuration is characteristic of glycoside hydrolase (GH) family 79 proteins. A structural similarity search showed that FoBGlcA mostly resembled GH79 β‐d‐glucuronidase (AcGlcA79A) of Acidobacterium capsulatum; however, the root‐mean‐square deviation value was 3.2 Å, indicating that FoBGlcA has a high structural divergence. FoBGlcA had a low sequence identity with AcGlcA79A (19%) and differed from other GH79 β‐glucuronidases. The structures of FoBGlcA and AcGlcA79A also differed in terms of the loop structure location near subsite –2 of their catalytic sites, which may account for the unique substrate specificity of FoBGlcA. The amino acid residues involved in the catalytic activity of this enzyme were determined by evaluating the activity levels of various mutant enzymes based on the crystal structure analysis of the FoBGlcA reaction product complex. Database Atomic coordinates and structure factors (codes https://doi.org/10.2210/pdb7DFQ/pdb and https://doi.org/10.2210/pdb7DFS/pdb) have been deposited in the Protein Data Bank (http://wwpdb.org/).

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Section: Resultsmentioning

confidence: 99%

Biochemical and structural characterization of a novel 4‐O‐α‐l‐rhamnosyl‐β‐d‐glucuronidase from Fusarium oxysporum

et al. 2021

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“…2015), whereas that of Bacillus subtilis were at E163 and E374 (Higuchi et al . 2018). Similarly, the conserved catalytic residues for GH‐42 have been reported to be at E142 and E304 in B. aryabhattai (Luan and Duan 2022), E142 and E312 in H. lacusprofundi (Laye et al .…”

Section: Classification Of β‐Galactosidasesmentioning

confidence: 99%

β‐Galactosidase: Insights into source variability, genetic engineering, immobilisation and diverse applications in food, industry and medicine

Zhou,

Liu,

Gao

et al. 2024

Int J of Dairy Tech

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β‐Galactosidases are crucial enzymes that hydrolyse oligosaccharides and polysaccharides with terminal β‐1,4‐glycosidic bonds. Though the traditional application of β‐Galactosidases has been to catalyse the breakdown of lactose in dairy products, its application extends beyond the production of lactose‐free products since variants capable of facilitating lactose condensation and exhibiting galactosyl transferase activity are extensively utilised for the synthesis of prebiotic galacto‐oligosaccharides. This review analyses β‐Galactosidase in multiple aspects, including sources, classification, characterisation, immobilisation, genetic engineering and applications in terms of whey treatment, biofuel production, production of lactose‐free dietary product, synthesis of galacto‐oligosaccharides and the early detection of cellular senescence and tumours.

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“…From the same research group, an enzyme was characterised as a 4,6Pyr-β- d -Gal-releasing enzyme (PyrGal-ase) with specificity for the (1→3) yeast linkage; mammalian (1→4)-linked PyrGal could not be hydrolysed. The physiological role of the PyrGal-ase in the Bacillus strain from where it was isolated is currently unknown [199].…”

Section: Ketal-pyruvyltransferasesmentioning

confidence: 99%

Pyruvate Substitutions on Glycoconjugates

Hager

Sützl

Stefanović

et al. 2019

IJMS

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Glycoconjugates are the most diverse biomolecules of life. Mostly located at the cell surface, they translate into cell-specific “barcodes” and offer a vast repertoire of functions, including support of cellular physiology, lifestyle, and pathogenicity. Functions can be fine-tuned by non-carbohydrate modifications on the constituting monosaccharides. Among these modifications is pyruvylation, which is present either in enol or ketal form. The most commonly best-understood example of pyruvylation is enol-pyruvylation of N-acetylglucosamine, which occurs at an early stage in the biosynthesis of the bacterial cell wall component peptidoglycan. Ketal-pyruvylation, in contrast, is present in diverse classes of glycoconjugates, from bacteria to algae to yeast—but not in humans. Mild purification strategies preventing the loss of the acid-labile ketal-pyruvyl group have led to a collection of elucidated pyruvylated glycan structures. However, knowledge of involved pyruvyltransferases creating a ring structure on various monosaccharides is scarce, mainly due to the lack of knowledge of fingerprint motifs of these enzymes and the unavailability of genome sequences of the organisms undergoing pyruvylation. This review compiles the current information on the widespread but under-investigated ketal-pyruvylation of monosaccharides, starting with different classes of pyruvylated glycoconjugates and associated functions, leading to pyruvyltransferases, their specificity and sequence space, and insight into pyruvate analytics.

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Identification and characterization of a novel β-D-galactosidase that releases pyruvylated galactose

Cited by 10 publications

References 31 publications

Biochemical and structural characterization of a novel 4‐O‐α‐l‐rhamnosyl‐β‐d‐glucuronidase from Fusarium oxysporum

Biochemical and structural characterization of a novel 4‐O‐α‐l‐rhamnosyl‐β‐d‐glucuronidase from Fusarium oxysporum

β‐Galactosidase: Insights into source variability, genetic engineering, immobilisation and diverse applications in food, industry and medicine

Pyruvate Substitutions on Glycoconjugates

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