A multidomain xylanase from a Bacillus sp. with a region homologous to thermostabilizing domains of thermophilic enzymes

Abstract

The gene xynC encoding xylanase C from Bacillus sp. BP-23 was cloned and expressed in Escherichia coli. The nucleotide sequence of a 3538 bp DNA fragment containing xynC gene was determined, revealing an open reading frame of 3258 bp that encodes a protein of 120567 Da. A comparison of the deduced amino acid sequence of xylanase C with known /I-glycanase sequences showed that the encoded enzyme is a modular protein containing three different domains. The central region of the enzyme is the catalytic domain, which shows high homology t o family 10 xylanases. A domain homologous t o family IX cellulose-binding domains is located in the C-terminal region of xylanase C, whilst the N-terminal region of the enzyme shows homology t o thermostabilizing domains found in several thermophilic enzymes. Xylanase C showed an activity profile similar t o that of enzymes from mesophilic microorganisms. Maximum activity was found at 45OC, and the enzyme was only stable at 55 O C or lower temperatures. Xylotetraose, xylotriose, xylobiose and xylose were the main products from birchwood xylan hydrolysis, whilst the enzyme showed increasing activity on xylo-oligosaccharides of increasing length, indicating that the cloned enzyme is an endoxylanase. A deletion derivative of xylanase C, lacking the region homologous t o thermostabilizing domains, was constructed. The truncated enzyme showed a lower optimum temperature for activity than the full-length enzyme, 35 O C instead of 4 5 OC, and a reduced thermal stability that resulted in a complete inactivation of the enzyme after 2 h incubation at 55 OC.

Microbial diversity in the larval gut of field and laboratory populations of the sugarcane weevil Sphenophorus levis (Coleoptera, Curculionidae)

“…Microorganisms from the Paenibacillus genus were isolated from laboratory larvae; this was the most abundant group of organisms to present CMCase activity. The strain Paenibacillus sp BP-23 (formerly Bacillus sp BP-23) (Blanco and Pastor, 1993) has been reported to have a multi-enzymatic glucanase system, including several cellulases (Pastor et al, 2001) and xylanases (Blanco et al, 1999). Several cellulolytic microorganisms were found in both populations, although the highest cellulolytic activity was observed in bacteria isolated from the laboratory population.…”

Biochemical and Structural Insights into Xylan Utilization by the Thermophilic Bacterium Caldanaerobius polysaccharolyticus

“…For aldouronic acids, the substrate (6 mg/ml) was incubated with 0.5 M Agu67A in 100 l of citrate buffer (50 4 ) was added for color development. The absorbance of the mixture was measured at 600 nm, and a standard curve was constructed using known concentrations of ␣-glucuronic acid (Sigma).…”

Characterization of a xylanase from a thermophilic strain of Anoxybacillus pushchinoensis A8

“…Among these enzymes, xylanases play a key role (Blanco et al 1999). The xylan backbone is degraded by β-1,4-xylanases and β-xylosidases.…”