anaerobic digester metagenome CE1 of multi-enzyme CE1-GH62-GH10 wtsFae1B
Comment
mutidomain enzyme consisting of three catalytic domains: ferulic acid esterase, arabinofuranosidase, and xylanase CE1-GH62-GH10. Here only N-terminal carbohydrate binding module family 48 and ferulic acid esterase (CE1) from the multi-enzyme CE1-GH62-GH10
(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > unclassified sequences: NE > metagenomes: NE > ecological metagenomes: NE > anaerobic digester metagenome: NE
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA QVTPRPEAAPGARPGFRAPARIISPEIMPDNKVTFRVYSKDASKVTITGE WQTGPGGVEELVKNDTGMFSITVGPLKPELYAYNFTVDGVKALDANNVQV RRDGTNYQNFFIIPGPESDLYFHKNNVPHGTVTKVWYKSSVIGFDRRMYV YTPAGYEGDTQRYPVFYLLHGAGGDEDAWTNMGRTAQIMDNLIAQGKAKP MIVVMTNGNANQAGAQNEVPPVPVTQGQQGIPSGSGMTGKFEEHLVKDVV PFIEKNFRALTGKDNRAIAGLSMGGGHTQTITNDNPGMFSYIGVFSMGIM AGRQQGADAEKIEKERDAKIEALKKSGYKLYWIACGKDDFVYQSALTLRN TLDKHNFKYVYRESTGGHTWANWRIYLSEFAPMLFK
References
Title: Enzymatic Cleavage of Diferuloyl Cross-Links in Corn Bran Arabinoxylan by Two Bacterial Feruloyl Esterases Lin S, Brask J, Munk L, Holck J, KBRM, Meyer AS, Agger JW, Wilkens C Ref: Journal of Agricultural and Food Chemistry, 70:133349, 2022 : PubMed
Corn bran is an abundant coprocessing stream of corn-starch processing, rich in highly substituted, diferuloyl-cross-linked glucurono-arabinoxylan. The diferuloyl cross-links make the glucurono-arabinoxylan recalcitrant to enzymatic conversion and constitute a hindrance for designing selective enzymatic upgrading of corn glucurono-arabinoxylan. Here, we show that two bacterial feruloyl esterases, wtsFae1A and wtsFae1B, each having a carbohydrate-binding module of family 48, are capable of cleaving the ester bonds of the cross-linkages and releasing 5-5', 8-5', 8-5' benzofuran, and 8-O-4' diferulate from soluble and insoluble corn bran glucurono-arabinoxylan. All four diferulic acids were released at similar efficiency, indicating nondiscriminatory enzymatic selectivity for the esterified dimer linkages, the only exception being that wtsFae1B had a surprisingly high propensity for releasing the dimers, especially 8-5' benzofuran diferulate, indicating a potential, unique catalytic selectivity. The data provide evidence of direct enzymatic release of diferulic acids from corn bran by newly discovered feruloyl esterases, i.e., a new enzyme activity. The findings yield new insight and create new opportunities for enzymatic opening of diferuloyl cross-linkages to pave the way for upgrading of recalcitrant arabinoxylans.
Feruloyl esterases (EC 3.1.1.73), belonging to carbohydrate esterase family 1 (CE1), hydrolyze ester bonds between ferulic acid (FA) and arabinose moieties in arabinoxylans. Recently, some CE1 enzymes identified in metagenomics studies have been predicted to contain a family 48 carbohydrate-binding module (CBM48), a CBM family associated with starch binding. Two of these CE1s, wastewater treatment sludge (wts) Fae1A and wtsFae1B isolated from wastewater treatment surplus sludge, have a cognate CBM48 domain and are feruloyl esterases, and wtsFae1A binds arabinoxylan. Here, we show that wtsFae1B also binds to arabinoxylan and that neither binds starch. Surface plasmon resonance analysis revealed that wtsFae1B's Kd for xylohexaose is 14.8 mum and that it does not bind to starch mimics, beta-cyclodextrin, or maltohexaose. Interestingly, in the absence of CBM48 domains, the CE1 regions from wtsFae1A and wtsFae1B did not bind arabinoxylan and were also unable to catalyze FA release from arabinoxylan. Pretreatment with a beta-d-1,4-xylanase did enable CE1 domain-mediated FA release from arabinoxylan in the absence of CBM48, indicating that CBM48 is essential for the CE1 activity on the polysaccharide. Crystal structures of wtsFae1A (at 1.63 A resolution) and wtsFae1B (1.98 A) revealed that both are folded proteins comprising structurally-conserved hydrogen bonds that lock the CBM48 position relative to that of the CE1 domain. wtsFae1A docking indicated that both enzymes accommodate the arabinoxylan backbone in a cleft at the CE1-CBM48 domain interface. Binding at this cleft appears to enable CE1 activities on polymeric arabinoxylan, illustrating an unexpected and crucial role of CBM48 domains for accommodating arabinoxylan.
anaerobic digester metagenome CE1 of multi-enzyme CE1-GH62-GH10 wtsFae1B
