The fructanolytic abilities of the rumen bacteriumButyrivibrio fibrisolvensstrain 3071

Abstract

Aims To determine if Butyrivibrio fibrisolvens strain 3071 is able to use fructose polymers for growth and to identify the enzymes involved in their digestion. Methods and Results Strain 3071 utilized 97, 89, 85 and 60% of sucrose, timothy grass fructan, inulin oligosaccharides and inulin, respectively, in the growth medium. A cell extract from timothy grass fructan‐grown bacteria was used for identification of fructanolytic enzymes by anion exchange chromatography, gel filtration, zymography and thin‐layer chromatography. The bacterium synthesizes a specific endolevanase and a nonspecific β‐fructofuranosidase. Both enzymes occurred in two forms differing in molecular weight. The β‐fructofuranosidase was not able to digest long‐chain inulin or timothy grass fructan, but degraded inulin oligosaccharides and sucrose. Addition of 1,4‐dithioerythritol to an enzyme solution did not affect the activity of endolevanase(s), but increased the ability of β‐fructofuranosidase to digest sucrose. The digestion of timothy grass fructan by endolevanase(s) was described by Michaelis–Menten kinetics in which Km = 2·82 g l−1 and Vmax = 4·01 μmoles reducing sugar equivalents × mg−1 × min−1. Conclusion Strain 3071 synthesizes enzymes enabling it to use grass fructans for growth. Significance and Impact of the Study Butyrivibrio fibrisolvens strain 3071 can be considered a member of the rumen fructanolytic guild.

A Highly Active Endo-Levanase BT1760 of a Dominant Mammalian Gut Commensal Bacteroides thetaiotaomicron Cleaves Not Only Various Bacterial Levans, but Also Levan of Timothy Grass

“…For example the identity of the endo-levanase LevB1 of Bacillus licheniformis [34] with BT1760 was 21.3% and that of the endo-levanase of Bacillus subtilis 168 [55] even lower–only 18.3%. An endo-levanase of a rumen bacterium Butyrivibrio fibrisolvens 3071 has also been biochemically characterized [25], but respective gene and protein sequences are not available. We then used the BLASTP program at NCBI (https://www.ncbi.nlm.nih.gov/) to search potential endo-levanase proteins in a human fecal isolate Butyrivibrio fibrisolvens 16/4 [56].…”

“…However, the activity was highest at pH 5.5–6.0 which was also confirmed in a levan degradation assay (S3 Fig). According to the literature, the endo-levanases from Bacillus licheniformis [34] and Butyrivibrio fibrisolvens [25] were assayed at pH 6.0 and the Bacillus sp. endo-levanase at pH 5.5 [66].…”

“…[25]. They showed that the Butyrivibrio fibrisolvens enzyme degrades timothy grass levan with maximum acivity (V max ) of 4 U/mg which is comparable to the rate of hydrolysis of LMW bacterial levan by endo-levanases of bacilli [34].…”

“…Our data show that the maximum velocity of BT1760 on timothy grass levan is over two hundred times higher indicating much more efficient catalysis. Still, the Butyrivibrio fibrisolvens endo-levanase has a good affinity to timothy grass levan, 2.82 g/L [25], that should contribute to efficient degradation of this fructan in the rumen.…”

“…For example, timothy grass ( Phleum pratense ) and orchard grass ( Dactylis glomerata ) have linear β-2,6-linked fructans which are referred to as phleins or plant levans [23–25]. Mixed levans (graminans) which contain both, β-2,1 and β-2,6 linkages, are present in many Poales species (e.g.…”