An Exopectate Lyase of Butyrivibrio fibrisolvens from the Bovine Rumen

Wojciechowicz, Maria; Heinrichová, Kveta; Ziołecki, A.

doi:10.1099/00221287-128-11-2661

Cited by 16 publications

(13 citation statements)

References 7 publications

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“…B. fibrisolvens also contains an exopectate lyase that generates unsaturated trigalacturonates, similar to PelA. To our knowledge, T. maritima and B. fibrisolvens are the only two bacteria described that contain such a similar combination of pectinolytic enzymes, although the exopolygalacturonase from B. fibrisolvens was shown to degrade both saturated and Δ4,5 unsaturated oligoGal p A [33].…”

Section: Discussionmentioning

confidence: 99%

Characterization and mode of action of an exopolygalacturonase from the hyperthermophilic bacterium Thermotoga maritima

et al. 2005

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Pectin is a complex polysaccharide present in the cell wall of higher plants, where it forms a network by embedding the other cell wall polysaccharides cellulose and hemicellulose. The backbone of the pectin molecule mainly consists of (partly methylated) homogalacturonan, interspersed with rhamnogalacturonan units, which often contain sugar side chains composed of arabinan and galactan [1].Degradation of the pectin polymer occurs via a set of pectinolytic enzymes, which can roughly be divided into esterases, which remove ferulic acid, methyl or acetyl groups, and depolymerases. The latter can be classified into lyases (b-elimination) and hydrolases [2]. All hydrolases involved in degradation of pectin are classified as members of family 28 of the glycoside hydrolases, including the endopolygalacturonases, exopolygalacturonases and rhamnogalacturonases [3,4]. Although a handful of endopolygalacturonases, generally of fungal origin [5][6][7][8][9][10], and a single rhamnogalacturonase [11] An intracellular pectinolytic enzyme, PelB (TM0437), from the hyperthermophilic bacterium Thermotoga maritima was functionally produced in Escherichia coli and purified to homogeneity. PelB belongs to family 28 of the glycoside hydrolases, consisting of pectin-hydrolysing enzymes. As one of the few bacterial exopolygalacturonases, it is able to remove monogalacturonate units from the nonreducing end of polygalacturonate. Detailed characterization of the enzyme showed that PelB is highly thermo-active and thermostable, with a melting temperature of 105°C and a temperature optimum of 80°C, the highest described to date for hydrolytic pectinases. PelB showed increasing activity on oligosaccharides with an increasing degree of polymerization. The highest activity was found on the pentamer (1000 UAEmg ). In addition, the affinity increased in conjunction with the length of the oligoGalpA chain. PelB displayed specificity for saturated oligoGalpA and was unable to degrade unsaturated or methyl-esterified oligoGalpA. Analogous to the exopolygalacturonase from Aspergillus tubingensis, it showed low activity with xylogalacturonan. Calculations on the subsite affinity revealed the presence of four subsites and a high affinity for GalpA at subsite +1, which is typical of exo-active enzymes. The physiological role of PelB and the previously characterized exopectate lyase PelA is discussed.Abbreviations PelB, exopolygalacturonase B; PelA, exopectate lyase A; PGA, polygalacturonic acid; (GalpA) n , oligogalacturonate with degree of polymerization n; DP, degree of polymerization; HPSEC, high-performance size-exclusion chromatography; HPAEC, high-performance anion-exchange chromatography.

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Section: Discussionmentioning

confidence: 99%

Characterization and mode of action of an exopolygalacturonase from the hyperthermophilic bacterium Thermotoga maritima

et al. 2005

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“…In particular, strains of Butyrivibrio fibrisolvens have been described that are cellulolytic (1,21,51), xylanolytic (12,23), amylolytic (10), pectinolytic (68), lipolytic (20), and proteolytic (11,54). This ability to ferment a wide range of macromolecules is considered to enable B. fibrisolvens to be the most dominant rumen bacterium under adverse nutritional conditions.…”

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confidence: 99%

Cloning, nucleotide sequence, and enzymatic characterization of an alpha-amylase from the ruminal bacterium Butyrivibrio fibrisolvens H17c

et al. 1991

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A Butyrivibrio fibrisolvens amylase gene was cloned and expressed by using its own promoter on the recombinant plasmid pBAMY100 in Escherichia coli. The amylase gene consisted of an open reading frame of 2,931 bp encoding a protein of 976 amino acids with a calculated Mr of 106,964. In E. coli(pBAMY100), more than 86% of the active amylase was located in the periplasm, and TnphoA fusion experiments showed that the enzyme had a functional signal peptide. The B. fibrisolvens amylase is a calcium metalloenzyme, and three conserved putative calcium-binding residues were identified. The amylase showed high sequence homology with other a-amylases in the three highly conserved regions which constitute the active centers. These and other conserved regions were located in the N-terminal half, and no similarity with any other amylase was detected in the remainder of the protein. Deletion of approximately 40% of the C-terminal portion of the amylase did not result in loss of amylolytic activity. The B. fibrisolvens amylase was identified as an endo-oa-amylase by hydrolysis of the Phadebas amylase substrate, hydrolysis of y-cyclodextrin to maltotriose, maltose, and glucose and the characteristic shape of the blue value and reducing sugar curves. Maltotriose was the major initial hydrolysis product from starch, although extended incubation resulted in its hydrolysis to maltose and glucose.Members of the genus Butyrivibrio are among the most numerous and nutritionally versatile of rumen bacteria. In particular, strains of Butyrivibrio fibrisolvens have been described that are cellulolytic (1, 21, 51), xylanolytic (12, 23), amylolytic (10), pectinolytic (68), lipolytic (20), and proteolytic (11,54). This ability to ferment a wide range of macromolecules is considered to enable B. fibrisolvens to be the most dominant rumen bacterium under adverse nutritional conditions. In winter, B. fibrisolvens is the most abundant starch-and cellulose-fermenting bacterium in the high-arctic Svalbard reindeer (43) and is unusual in its capacity to utilize both carbohydrate substrates whereas other cellulolytic species are unable to do so. B. fibrisolvens is also the most prevalent bacterium in the rumen of semistarved Zebu cattle in Kenya (35).Starch is an important component of the ruminal diet, and its digestion is essential for maximum productivity. Highstarch diets are, however, associated with a number of digestive disorders. For example, lactate acidosis is associated with a proliferation of the amylolytic species Streptococcus bovis and is thought to occur as a result of the rapid fermentation of starch leading to an accumulation of lactic acid (53). A knowledge of the nature and regulation of amylolytic enzymes could therefore assist in controlling these digestive disorders and improving the efficiency of starch digestion. Relatively few amylases of ruminal bacteria have been characterized. Those that have been studied include amylases from S. bovis, Clostridium butyricum (24, 65), and Ruminobacter amylophilis (formerly Bacteroides am...

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“…Since then, amylolytic, xylanolytic and even pectinolytic properties of strains belonging to this species have been documented in numerous studies (Wojciechowicz et al . ; Stewart et al . ).…”

Section: Introductionmentioning

confidence: 99%

“…The species Butyrivibrio fibrisolvens was described by Bryant and Small (1956) as a butyrate-producing curved rod-shaped rumen bacteria able to digest and then to ferment the majority of polysaccharides occurring in ruminant diets, i.e., dextrin, starch, xylan and in minor amounts, cellulose. Since then, amylolytic, xylanolytic and even pectinolytic properties of strains belonging to this species have been documented in numerous studies (Wojciechowicz et al 1982;Stewart et al 1997).…”

Section: Introductionmentioning

confidence: 99%

The fructanolytic abilities of the rumen bacteriumButyrivibrio fibrisolvensstrain 3071

et al. 2015

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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.

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An Exopectate Lyase of Butyrivibrio fibrisolvens from the Bovine Rumen

Cited by 16 publications

References 7 publications

Characterization and mode of action of an exopolygalacturonase from the hyperthermophilic bacterium Thermotoga maritima

Characterization and mode of action of an exopolygalacturonase from the hyperthermophilic bacterium Thermotoga maritima

Cloning, nucleotide sequence, and enzymatic characterization of an alpha-amylase from the ruminal bacterium Butyrivibrio fibrisolvens H17c

The fructanolytic abilities of the rumen bacteriumButyrivibrio fibrisolvensstrain 3071

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