Dataset Information

Screening for and identification of starch-, amylopectin-, and pullulan-degrading activities in bifidobacterial strains.

ABSTRACT: Forty-two bifidobacterial strains were screened for alpha-amylase and/or pullulanase activity by investigating their capacities to utilize starch, amylopectin, or pullulan. Of the 42 bifidobacterial strains tested, 19 were capable of degrading potato starch. Of these 19 strains, 11 were able to degrade starch and amylopectin, as well as pullulan. These 11 strains, which were shown to produce extracellular starch-degrading activities, included 5 strains of Bifidobacterium breve, 1 B. dentium strain, 1 B. infantis strain, 3 strains of B. pseudolongum, and 1 strain of B. thermophilum. Quantitative and qualitative enzyme activities were determined by measuring the concentrations of released reducing sugars and by high-performance thin-layer chromatography, respectively. These analyses confirmed both the inducible nature and the extracellular nature of the starch- and pullulan-degrading enzyme activities and showed that the five B. breve strains produced an activity that is consistent with type II pullulanase (amylopullulanase) activity, while the remaining six strains produced an activity with properties that resemble those of type III pullulan hydrolase.

SUBMITTER: Ryan SM

PROVIDER: S-EPMC1538741 | biostudies-literature | 2006 Aug

REPOSITORIES: biostudies-literature

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Screening for and identification of starch-, amylopectin-, and pullulan-degrading activities in bifidobacterial strains.

Ryan Sinéad M SM Fitzgerald Gerald F GF van Sinderen Douwe D

Applied and environmental microbiology 20060801 8

Forty-two bifidobacterial strains were screened for alpha-amylase and/or pullulanase activity by investigating their capacities to utilize starch, amylopectin, or pullulan. Of the 42 bifidobacterial strains tested, 19 were capable of degrading potato starch. Of these 19 strains, 11 were able to degrade starch and amylopectin, as well as pullulan. These 11 strains, which were shown to produce extracellular starch-degrading activities, included 5 strains of Bifidobacterium breve, 1 B. dentium stra ...[more]

PMID: 16885278

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Project description:BackgroundThe biochemical mechanisms that determine the molecular architecture of amylopectin are central in plant biology because they allow long-term storage of reduced carbon. Amylopectin structure imparts the ability to form semi-crystalline starch granules, which in turn provides its glucose storage function. The enzymatic steps of amylopectin biosynthesis resemble those of the soluble polymer glycogen, however, the reasons for amylopectin's architectural distinctions are not clearly understood. The multiplicity of starch biosynthetic enzymes conserved in plants likely is involved. For example, amylopectin chain elongation in plants involves five conserved classes of starch synthase (SS), whereas glycogen biosynthesis typically requires only one class of glycogen synthase.ResultsNull mutations were characterized in AtSS2, which codes for SSII, and mutant lines were compared to lines lacking SSIII and to an Atss2, Atss3 double mutant. Loss of SSII did not affect growth rate or starch quantity, but caused increased amylose/amylopectin ratio, increased total amylose, and deficiency in amylopectin chains with degree of polymerization (DP) 12 to DP28. In contrast, loss of both SSII and SSIII caused slower plant growth and dramatically reduced starch content. Extreme deficiency in DP12 to DP28 chains occurred in the double mutant, far more severe than the summed changes in SSII- or SSIII-deficient plants lacking only one of the two enzymes.ConclusionSSII and SSIII have partially redundant functions in determination of amylopectin structure, and these roles cannot be substituted by any other conserved SS, specifically SSI, GBSSI, or SSIV. Even though SSIII is not required for the normal abundance of glucan chains of DP12 to DP18, the enzyme clearly is capable of functioning in production such chains. The role of SSIII in producing these chains cannot be detected simply by analysis of an individual mutation. Competition between different SSs for binding to substrate could in part explain the specific distribution of glucan chains within amylopectin.

Project description:Probiotics might offer an attractive alternative to prevent and control Clostridium difficile (C. difficile) infection (CDI). Limited information is available on the ability of commercially used bifidobacterial strains to inhibit C. difficile. This study examined the anti-clostridial effects of Bifidobacterium longum JDM301, a widely used commercial probiotic strain in China, in vitro and in vivo. In vitro evaluation revealed a significant reduction in C. difficile counts when JDM301 was co-cultured with C. difficile, which was correlated with the significant decrease in clostridial toxin titres (TcdA and TcdB). Furthermore, the cell-free culture supernatants (CFS) of JDM301 inhibited C. difficile growth and degraded TcdA and TcdB. Notably, the results showed that acid pH promoted the degradation of TcdA by CFS from JDM301. Furthermore, comparative studies among 10 B. longum strains were performed, which showed that the inhibitory effect of CFS from JDM301 was similar with the other 8 B. longum strains and higher than strain BLY1. However, when it was neutralized, the significant different was lost. When present together, it was suggested that the acid pH induced by probiotics not only played important roles in the growth inhibition against C. difficile resulting in the reduction of toxins titres, but also directly promoted the degradation of clostridial toxin. In vivo studies proved that JDM301 partially relieved damage to tissues caused by C. difficile and also decreased the number of C. difficile and toxin levels. In summary, our results demonstrated that the commercial strain, JDM301 could be considered a probiotic able to exert anti-toxin capability and most of the CFS from Bifidobacterium were able to inhibit the growth of C. difficile, depending on acid pH. These results highlighted a potential that JDM301 could be helpful in preventing CDI and that most of the bifidobacterial strains could (at least partially) exert protective effects by reducing toxin titres through growth inhibition against toxigenic C. difficile.

Project description:Three starch branching enzyme (BE) isozymes, BEI, BEIIa, and BEIIb, are involved in starch biosynthesis in rice endosperm. Past in vivo and in vitro studies have suggested that each BE isozyme plays a distinct role in forming the fine structure of amylopectin. To elucidate more details of their roles, we prepared DNA constructs in which all the possible combinations of the expressions of these three isozymes were suppressed in developing rice endosperm. Analysis of the chain-length distributions of amylopectin produced under these various conditions confirmed the contributions of the individual BE isozymes to the fine structure of amylopectin in rice endosperm. Among these isozymes, the impact of loss of BEIIb activity on amylopectin fine structure was most remarkable and indicated that it plays a specific role in the synthesis of short chains with a 6-13 degree of polymerization (DP). The contribution of BEI to the amylopectin synthesis was unclear when only BEI activity was reduced. It was clear, however, when both BEI and BEIIb activities were substantially inhibited. The DP11-22 intermediate chains were markedly reduced in the ΔBEI/BEIIb line compared with the ΔBEIIb line, indicating that BEI plays a distinct role in the synthesis of these intermediate chains. Although no substantial change in amylopectin chain profile was detected in the ΔBEIIa line, the role of BEIIa could be deciphered by analyzing amylopectin fine structure from the ΔBEI/BEIIa/BEIIb line in comparison to that from ΔBEI/BEIIb line. This strongly suggests that BEIIa compensates for the role of BEI, rather than that of BEIIb, by forming intermediate chains of DP11-22. In addition, the new possibility that BEIIa is involved in the formation of starch granules in rice endosperm was suggested because the onset temperature for gelatinization of starch granules in the ΔBEIIa/BEIIb line was significantly higher than that in the ΔBEIIb line. In summary, the present study highlights the distinct roles of BEI, BEIIa, and BEIIb in the synthesis of amylopectin in developing rice endosperm.

Dataset Information

Screening for and identification of starch-, amylopectin-, and pullulan-degrading activities in bifidobacterial strains.

Publications

Screening for and identification of starch-, amylopectin-, and pullulan-degrading activities in bifidobacterial strains.

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