Isolation of Clostridium acetobutylicum strains and the preliminary investigation of the hemicellulolytic activities of isolate 3BYR

Abstract

Thirty-nine xylanolytic bacteria tentatively identified as Clostridia were isolated from a selection of agricultural and forest soil samples, and water–sediment–fibre samples from acidic springs in the Yellowstone National Park. Screening for xylan hydrolysis was performed using an enriched agar medium at pH 5.5 with Remazol–xylan as the indicator, with or without an initial enrichment using xylan as the major carbon source. From 13 of the most highly xylanolytic strains, 9 were tentatively identified as Clostridium acetobutylicum, 2 as Clostridium butyricum, and 2 as Clostridium beijerinckii. The C. acetobutylicum isolate 3BYR utilized 80% of oat spelt xylan as a carbon source during growth. The bacterium exhibited very high extracellular xylanase and xylosidase activities and, as well, α-L-arabinofuranosidase and α-glucuronidase activities. Glucuronidase activity was documented by the release of 4-O-methyl-α-D-glucuronic acid from birchwood xylan. The results of this work indicate the ubiquity of xylanolytic Clostridia, and that the previously unreported activity, α-glucuronidase, has been demonstrated in C. acetobutylicum. Key words: α-glucuronidase, xylanase, Clostridium acetobutylicum, xylan.

Isolation and analysis of a gene encoding α‐glucuronidase, an enzyme with a novel primary structure involved in the breakdown of xylan

“…This enzyme activity was also reported to occur in some bacteria, i.e. Streptomyces olivochromogenes (MacKenzie et al, 1987), Fibrobacter succinogenes (Smith and Forsberg, 1991), Clostridium acetobutylicum (Trudeau et al, 1992), Clostridium stercorarium (Bronnenmeier et al, 1995b), Thermoanaerobacterium saccharolyticum (Bronnenmeier et al, 1995b) and Thermoanaerobacterium sp. strain JW/SL-YS485 (Shao et al, 1995).…”

“…Alpha-D-glucuronidase activity has been detected in a variety of fungi and in some bacteria, and several enzymes have been purified (Bronnenmeier et al, 1995b;Ishihara et al, 1990;Khandke et al, 1989;MacKenzie et al, 1987;Puls, 1992;Shao et al, 1995;Smith and Forsberg, 1991;Trudeau et al, 1992;Uchida et al, 1992). To date, however, no primary structures of ␣-glucuronidases, whether of fungal or bacterial origin, are known and we are not aware of any reports describing the isolation of ␣-glucuronidase genes.…”

A Chromogenic Substrate for a β-Xylosidase-Coupled Assay of α-Glucuronidase

“…A nonmodified procedure for determining reducing sugars was used in the case of aldouronic acids reduced with NaBH 4 (9). Alternative methods include HPLC determination of free MeGlcA (19,20) or the equivalent amount of xylooligosaccharide (12,21), or GLC of its trimethylsilyl oxim derivative (7,(22)(23)(24).…”