Three-dimensional structures and functional studies of two GH43 arabinofuranosidases fromWeissellasp. strain 142 andLactobacillus brevis

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Abstract

Arabinofuranosidases degrade arabinose-containing oligo and polysaccharides, releasing l-arabinose, which is a potentially useful sugar, shown to reduce glycemic response under certain conditions. Arabinofuranosidases (Arafs) are frequently found in GH43, one of the most common GH-families encoded in genomes in gut microbiota, and hence it is of interest to increase understanding of the function of these enzymes in species occurring in the gut. Here we have produced, characterized and solved the three-dimensional structures, at 1.9 and 2.0 Å resolution respectively, of two homologous GH43 enzymes, classified under subfamily 26, from Lactobacillus brevis DSM1269 (LbAraf43) and Weissella strain 142 (WAraf43), respectively. The enzymes, with 74% sequence identity to each other, are composed of a single catalytic module with a β-propeller structure typical of GH43, and an active-site pocket with three identifiable subsites (−1, +1, and +2). According to size exclusion chromatography, native WAraf43 is a dimer, while LbAraf43 is a tetramer in solution. Both of them show activity with similar catalytic efficiency on 1,5-α-l-arabinooligosaccharides with a degree of polymerization (DP) of 2–3. Activity is restricted to substrates of low DP, and the reason for this is believed to be an extended loop at the entrance to the active site, creating interactions in the +2 subsite.

Database

Structural data are available in the PDB under the accession numbers 5M8B (LbAraf43) and 5M8E (WAraf43).

Three-dimensional structures of GH43 arabinofuranosidases, from Lactobacillus brevis DSM1269 and Weissella strain 142 were solved and displayed single-domain β-propeller structures with a metal ion coordination site located behind the catalytic pocket. Activity was restricted to arabinooligosaccharides of low DP (2-3), which may be due to the extended loop in blade II (red) at the entrance to the active site.

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