Cooperation between β‐galactosidase and an isoprimeverose‐producing oligoxyloglucan hydrolase is key for xyloglucan degradation in Aspergillus oryzae

Abstract

The galactosylation of xyloglucan blocks many of the enzymatic processes targeting this oligosaccharide. We found that the expression of a gene encoding Aspergillus oryzae β‐galactosidase (LacA) is induced in the presence of xyloglucan oligosaccharides. With detailed analyses of the substrate specificity of purified recombinant LacA, we show that LacA cleaves galactopyranosyl residues from xyloglucan oligosaccharides, but not from xyloglucan polysaccharide, and plays a vital role in xyloglucan degradation. LacA acts cooperatively with the isoprimeverose‐producing oligoxyloglucan hydrolase IpeA to hydrolyze xyloglucan oligosaccharides. Galactosylation of the xylopyranosyl side chain at the nonreducing end of oligoxyloglucan saccharides completely abolishes IpeA activity while LacA efficiently removes the galactopyranosyl residue. Conversely, an isoprimeverose unit at the nonreducing end of the main chain of xyloglucan oligosaccharides blocks LacA activity, while IpeA can still remove the isoprimeverose moiety. This is the first study reporting the cooperative action of β‐galactosidase and isoprimeverose‐producing oligoxyloglucan hydrolase on xyloglucan oligosaccharide degradation. Our findings shed light on the true role of LacA and the enzymatic coordination between β‐galactosidase and other hydrolases on xyloglucan degradation.

Complementary Contribution of Fungi and Bacteria to Lignocellulose Digestion in the Food Stored by a Neotropical Higher Termite

“…On the other hand, lignocellulases in the gut of C. cumulans were linked to Firmicutes, Ascomycota, and Chytridiomycota. All these microbes dominated the core microbiota and are known to produce cellulases, hemicellulases (Couturier et al, 2016;Grieco et al, 2019;Matsuzawa et al, 2019Matsuzawa et al, , 2020 and laccases (Murphy et al, 2021). In fungus-growing insects that exploit plant-derived resources, the association between fungi and bacteria result in a multipartite functional metabolism of lignocellulose (Barcoto et al, 2020).…”

Identification and Characterization of Novel Intracellular α-Xylosidase in <i>Aspergillus oryzae</i>

“…Aspergillus oryzae has been used to produce traditional Japanese fermented foods such as miso, soy sauce, and Japanese rice wine (sake). A. oryzae produces many polysaccharide degradation-related enzymes 8) 9) including xyloglucan degradation-related enzymes such as two xyloglucan-specific endo-β-1,4-glucanases (xyloglucanases, Xeg5A and Xeg12A, EC 3.2.1.151), 10) isoprimeverose-producing oligoxyloglucan hydrolase (IpeA, EC 3.2.1.120), 11) 12) β-galactosidase (LacA, EC 3.2.1.23), 13) and two α-xylosidases (AxyA and AxyB, EC 3.2.1.177). 14) 15) Xeg5A and Xeg12A degrade xyloglucans into xyloglucan oligosaccharides.…”

“…IpeA releases isoprimeverose [α-D-xylopyranosyl-(1→6)-D-glucose] from the non-reducing ends of xyloglucan oligosaccharides, 16) 17) whereas LacA releases galactose from the side chains of xyloglucan oligosaccharides. 13) Isoprimeverose is then hydrolyzed to D-xylose and D-glucose by AxyA and AxyB. 14) 15) AxyB contains an N-terminal signal sequence for its secretion.…”

Structural basis for the catalytic mechanism of the glycoside hydrolase family 3 isoprimeverose‐producing oligoxyloglucan hydrolase from Aspergillus oryzae

“…These xyloglucanases belong to the glycoside hydrolase (GH) family 5 and GH12, respectively, and degrade xyloglucan polysaccharides into xyloglucan oligosaccharides through different modes of action [9]. Xyloglucan oligosaccharides are then degraded into diand monosaccharides by exo-type enzymes such as isoprimeverose-producing oligoxyloglucan hydrolase (IpeA), b-galactosidase (LacA), and a-xylosidases (AxyA and AxyB) [10][11][12][13]. IpeA, LacA, and AxyA/B belong to GH3, GH35, and GH31, respectively, and D-xylose and xyloglucan oligosaccharides induce the expression of genes encoding these enzymes.…”

“…Although most GH3 enzymes recognize and release monosaccharides from the non-reducing end of di-and oligosaccharides, IpeA strictly recognizes and releases an isoprimeverose unit (a-D-xylopyranosyl-(1?6)-b-Dglucopyranose) from the non-reducing end of the b-1,4-glucan main chain of xyloglucan oligosaccharides [10]. As xylopyranosyl side-chain galactosylation of xyloglucan oligosaccharides blocks IpeA activity, the cooperation of IpeA and LacA is essential for xyloglucan degradation [11]. AxyA and AxyB then degrade the released isoprimeverose into D-xylose and Dglucose [12,13].…”