Characterization of a GH Family 43 β-Xylosidase Having a Novel Carbohydrate-binding Module from <i>Paenibacillus xylaniclasticus</i> Strain TW1

Abstract

Paenibacillus xylaniclasticus strain TW1, a gram-positive facultative anaerobic bacterium, was isolated as a xylanolytic microorganism from the wastes of a pineapple processing factory. A gene encoding one of its xylanolytic enzymes, a β-xylosidase, was cloned and sequenced. Sequence analysis revealed that this β-xylosidase, named PxXyl43A, was composed of a glycoside hydrolase (GH) family 43 subfamily 12 catalytic module and an unknown function module (UM). The full-length PxXyl43A (PxXyl43A) was heterologously expressed in Escherichia coli and purified. Recombinant PxXyl43A exhibited hydrolysis activity against both p-nitrophenyl-β-D-xylopyranoside (pNPX) and p-nitrophenyl-α-L-arabinofuranoside at specific activities of 250 and 310 mU/mg, respectively. The optimal reaction pH and temperature for pNPX hydrolysis were 7.1 and 54 ˚C, respectively. At pH 7.0 and 54 ˚C, the Km and kcat for pNPX were 1.2 mM and 2.8 0.15 s -1 , respectively. It was also discovered that the recombinant unknown function module of PxXyl43A (PxXyl43A-UM) could bind to insoluble xylans like birchwood xylan and oat spelt xylan, whereas it did not bind to cellulosic substrates such as ball-milled cellulose, carboxymethyl cellulose or lichenan. The PxXyl43A-UMʼs binding constant value Ka for oat spelt xylan was 2.0 10 -5 M -1 . These results suggest that PxXyl43A possesses a novel carbohydrate-binding module, named as CBM91, specific for xylan-containing polysaccharides.

A metagenomic survey of the fecal microbiome of the African savanna elephant (Loxodonta africana)

“…Among the 40 CBMs identified, CBM67, CBM13, CBM2, CBM91 and CBM20 were found to be present at a relative abundance of >1% (Table 6). This group of binding modules has a propensity to bind components of hemicellulose and cellulose, with CBM67 binding l ‐rhamnose (Fujimoto et al., 2013), CBM13 targeting xylose/xylan (Tomme et al., 2000), CBM91 specific to xylan (Ito et al., 2022) and CBM2 binding to cellulose (Kostylev et al., 2012). The exception in this group is CBM20, which is defined as a starch binding domain (Christiansen et al., 2009).…”

Diversity and host interaction of the gut microbiota in specific pathogen-free pigs

“…Interestingly, CBM91 exhibited lower abundance in the small intestine and stomach, but higher abundance in the large intestine and rectum (Supplementary Figure S6D). CBM91 is capable of binding xylan substances (such as birch and oat bran) (Ito et al, 2022), and the large intestine and rectum are the sites of fiber digestion (Moran and Bedford, 2022). The abundance of CBM50 gradually decreased along the GIT (Supplementary Figure S6D); CBM50 is involved in the degradation of chitin or peptidoglycans (Gruber et al, 2011).…”

A C1/C4-Oxidizing AA10 Lytic Polysaccharide Monooxygenase from <i>Paenibacillus xylaniclasticus</i> Strain TW1

“… 4) Paenibacillus xylaniclasticus strain TW1 is a Gram-positive and facultative bacterium that has many genes encoding cellulolytic/xylanolytic enzymes. 5) 6) P. xylaniclasticus TW1 has an LPMO gene encoding Px AA10A that belongs to the AA10 family. However, both C1/C4 selectivity for the oxidized position and the enzyme activity of Px AA10A remain to be characterized.…”