MOESM1 of Biochemical and structural features of diverse bacterial glucuronoyl esterases facilitating recalcitrant biomass conversion
Creators: name: Bååth, Jenny Arnling
name: Mazurkewich, Scott
name: Knudsen, Rasmus
name: Jens-Christian Poulsen
name: Olsson, Lisbeth
name: Leggio, Leila Lo
name: Larsbrink, Johan
Publisher: name: figshare
Year: 2018
Subjects: subject: Biochemistry
subject: Medicine
subject: Microbiology
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Genetics
subject: Evolutionary Biology
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: FOS: Chemical sciences
subject: Ecology
subject: 69999 Biological Sciences not elsewhere classified
subject: 110309 Infectious Diseases
subject: FOS: Health sciences
subject: FOS: Health sciences
subject: Plant Biology

description: Additional file 1: Table S1. Percent sequence identity and percent query coverage (in brackets) between all CE15 enzymes used in this study. Sequence identity values for CE15 enzymes within one organism are marked green (O. terrae), magenta (S. linguale) and blue (S. usitatus). The query sequences are presented in the top row.Kinetic parameters of O. terrae, S. linguale, and S. usitatus CE15 enzymes on model. Table S2. Kinetic parameters of O. terrae, S. linguale, and S. usitatus CE15 enzymes on model substrates. Esterase activity with benzyl (Bnz), allyl (Allyl), methyl (Me) esters of glucuronoate (GlcA) and galacturonoate (GalA) are shown in addition to acetyl esterase activity with 4-nitrophenol acetate (pNP-Ac) and 1,2,3,4-tetra-O-acetyl-β-d-xylopyranose (TetAcXyl). Table S3. Primers used for cloning CE15 constructs and for qPCR of S. linguale CE15 members. Table S4. Table of crystallographic statistics. Figure S1. Unrooted phylogenetic tree of all members of CE15 (catalytic domains), with Genbank accession numbers as identifiers. Yellow branches represent fungal members, circles indicate biochemically characterized members, and stars represent members with solved structures. Targets of this study are shown using the same color code as in the main text: green for O. terrae, red for S. linguale, and blue for S. usitatus. Figure S2. Model substrates used in this study: (A) BnzGlcA, (B) AllylGlcA, (C) MeGlcA, (D) MeGalA, (E) pNP-Ac and (F) TetAcXyl. Figure S3. Effect of pH on BnzGlcA esterase activity for CE15 enzymes from O. terrae (OtCE15 A-D, panels A-D), S. linguale (SlCE15 A-C, panels E-G), and S. usitatus (SuCE15 A-C, panels H-J). Mean values of relative activity from duplicate measurements are plotted with standard error of the mean. Figure S4. Structure-based sequence alignment of all CE15 enzymes structurally characterized to date. Similar residues are written in red text while conserved residues are written in white text over a red background. The insertion regions found in the bacterial structures relative to the fungal counterparts are highlighted in yellow. The residues of the canonical catalytic triad are indicated by cyan arrows below the text. The aspartate in MZ0003 proposed to act as the acidic residue of the catalytic triad, in place of the missing canonical glutamate, is indicate by a black arrow below the text. Note that both OtCE15A and SuCE15C also have an aspartate at the same position while additionally having the glutamate of the canonical catalytic triad. Residues hydrogen bonding with 4-O-methyl-glucuronoate in the StGE2 co-crystal structure are indicated by blue arrows above the text. The isoleucine and leucine comprising a hydrophobic patch near the 4-O-methyl substituent in the StGE2 co-crystal structure are indicated by magenta arrows. The phenylalanine conserved in the bacterial structures possibly aiding in positioning in aromatic substituents of the sugar esters is indicated with a grey arrow. The disulfide bridges formed in the fungal structures are indicated above the alignment by numbering in green text. Figure S5. Multiple sequence alignment of characterized glucuronoyl esterases. Similar residues are written in red text while conserved residues are written in white text over a red background. The insertion regions found in the bacterial structures relative to the fungal counterparts are highlighted in yellow. The residues of the conserved catalytic triad are colored cyan. Note that glutamate of the catalytic triad is not conserved in all bacterial esterases and the position of the equivalent acidic residue in MZ0003 is also colored cyan. Arrows indicating significant residues are colored as in Additional file 5: Figure S4. Figure S6. Growth curves of S. linguale when grown with different additives or on different carbon sources. S. linguale did not grow on standard minimal media and an optimized media for bacterial growth was determined experimentally (see methods for formulation). (A) Growth curves of S. linguale in the optimized media without a carbon source (red), with 0.3% (w/v) glucose (blue) and in the media containing glucose but in the absence of either trace metals and vitamins (green), sodium phosphate pH 7.5 (magenta), or magnesium sulphate (cyan). (B) Growth of S. linguale in optimized media with 0.3% of either glucose (blue), xylose (purple), or xylan from corn cob (yellow).