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emi2856-sup-0001-fS1.pdf28K

Fig. S1. HHpred alignment of UxaE from T. maritima (TM0440) and the highest scoring protein with experimentally determined structure (tagatose-bisphosphate aldolase AgaY, pdb entry 1gvf). Experimental and predicted secondary structure is shown above and below the sequences: H – helix; E – strand; C, T – coil and turn; small letters denote less confident prediction.

emi2856-sup-0002-fS2.pdf166K

Fig. S2. A. Phylogenetic tree of the UxaE family proteins. The tree was constructed using the maximum likelihood method (phyml) from the phylip package using the default parameters and visualized with Dendroscope. The numbers represent bootstrap values in percentages obtained from 1000 replicates.

B. Multiple alignment of UxaE family proteins analysed in this work. Conserved residues likely involved in the catalysis and discussed in the text are indicated.

emi2856-sup-0003-fS3.pdf20K

Fig. S3. GC-MS analysis of hexuronic acid isomerization by tm_UxaC. The assay mixture containing either d-GlcA (A) or d-GalA (B) (black line) was incubated overnight with the recombinant tm_UxaC enzyme and showed the appearance of new peaks that correspond to either d-FruA (A) or d-TagA (B) respectively (red line). Retention times of hexuronic acid-specific peaks as determined on total ion count chromatogram: d-GlcA, Rt = 10.44 and 10.57 min; d-FruA, Rt = 10.15 and 10.27 min; d-GalA, Rt = 10.52 and 10.68 min; d-TagA, Rt = 10.39 and 10.51 min; latter peak obscured by d-GalA.

emi2856-sup-0004-fS4.pdf96K

Fig. S4. Genetic complementation of E. coli ΔuxaB and ΔuxuB mutant strains by the pMH2T7-derived plasmids harbouring TM0442 (encoding tm_GntE), TM0440 (encoding tm_UxaE) or TM0808 gene (used as a negative control). The recombinant strains were spotted on M9 agar plates supplemented with 4 g l−1 either d-GalA (A), d-GlcA (B) or d-gluconate (C) (used as a positive control) and incubated at 37°C for 24 h.

emi2856-sup-0005-fS5.pdf61K

Fig. S5. Electron ionization mass spectra obtained for d-GlcA (A), d-GalA (B), d-FruA (C) and d-TagA (D). d-GalA and d-GlcA epimers have five characteristic m/z fragments (423, 393, 364, 333 and 305). d-TagA and d-FruA epimers are characterized by two abundant m/z fragments (364 and 277).

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