Dataset Information

Discovery of a New Microbial Origin Cold-Active Neopullulanase Capable for Effective Conversion of Pullulan to Panose.

ABSTRACT: Panose is a type of functional sugar with diverse bioactivities. The enzymatic conversion bioprocess to produce high purity panose with high efficiency has become increasingly important. Here, a new neopullulanase (NPase), Amy117 from B. pseudofirmus 703, was identified and characterized. Amy117 presented the optimal activity at pH 7.0 and 30 °C, its activity is over 40% at 10 °C and over 80% at 20 °C, which is cold-active. The enzyme cleaved α-1, 4-glycosidic linkages of pullulan to generate panose as the only hydrolysis product, and degraded cyclodextrins (CDs) and starch to glucose and maltose, with an apparent preference for CDs. Furthermore, Amy117 can produce 72.7 mg/mL panose with a conversion yield of 91% (w/w) based on 80 mg/mL pullulan. The sequence and structure analysis showed that the low proportion of Arg, high proportion of Asn and Gln, and high α-helix levels in Amy117 may contribute to its cold-active properties. Root mean square deviation (RMSD) analysis also showed that Amy117 is more flexible than two mesophilic homologues. Hence, we discovered a new high-efficiency panose-producing NPase, which so far achieves the highest panose production and would be an ideal candidate in the food industry.

SUBMITTER: Wang M

PROVIDER: S-EPMC9267027 | biostudies-literature | 2022 Jun

REPOSITORIES: biostudies-literature

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Discovery of a New Microbial Origin Cold-Active Neopullulanase Capable for Effective Conversion of Pullulan to Panose.

Wang Meixing M Hu Huizhen H Zhang Buyu B Zheng Yang Y Wu Pan P Lu Zhenghui Z Zhang Guimin G

International journal of molecular sciences 20220622 13

Panose is a type of functional sugar with diverse bioactivities. The enzymatic conversion bioprocess to produce high purity panose with high efficiency has become increasingly important. Here, a new neopullulanase (NPase), Amy117 from <i>B. pseudofirmus</i> 703, was identified and characterized. Amy117 presented the optimal activity at pH 7.0 and 30 °C, its activity is over 40% at 10 °C and over 80% at 20 °C, which is cold-active. The enzyme cleaved α-1, 4-glycosidic linkages of pullulan to gene ...[more]

PMID: 35805929

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Project description:Enriched microbial communities, obtained from environmental samples through selective processes, can effectively contribute to lignocellulose degradation. Unfortunately, fully controlled industrial degradation processes are difficult to reach given the intrinsically dynamic nature and complexity of the microbial communities, composed of a large number of culturable and unculturable species. The use of less complex but equally effective microbial consortia could improve their applications by allowing for more controlled industrial processes. Here, we combined ecological theory and enrichment principles to develop an effective lignocellulose-degrading minimal active microbial Consortia (MAMC). Following an enrichment of soil bacteria capable of degrading lignocellulose material from sugarcane origin, we applied a reductive-screening approach based on molecular phenotyping, identification, and metabolic characterization to obtain a selection of 18 lignocellulose-degrading strains representing four metabolic functional groups. We then generated 65 compositional replicates of MAMC containing five species each, which vary in the number of functional groups, metabolic potential, and degradation capacity. The characterization of the MAMC according to their degradation capacities and functional diversity measurements revealed that functional diversity positively correlated with the degradation of the most complex lignocellulosic fraction (lignin), indicating the importance of metabolic complementarity, whereas cellulose and hemicellulose degradation were either negatively or not affected by functional diversity. The screening method described here successfully led to the selection of effective MAMC, whose degradation potential reached up 96.5% of the degradation rates when all 18 species were present. A total of seven assembled synthetic communities were identified as the most effective MAMC. A consortium containing Stenotrophomonas maltophilia, Paenibacillus sp., Microbacterium sp., Chryseobacterium taiwanense, and Brevundimonas sp. was found to be the most effective degrading synthetic community.

Project description:BackgroundTemperate bacteriophages are capable of lysogenic conversion of new bacterial hosts. This phenomenon is often ascribed to "moron" elements that are acquired horizontally and transcribed independently from the rest of the phage genes. Whereas some bacterial species exhibit relatively little prophage-dependent phenotypic changes, other bacterial species such as Stenotrophomonas maltophilia appear to commonly adopt prophage genetic contributions.ResultsThe novel S. maltophilia bacteriophage DLP4 was isolated from soil using the highly antibiotic-resistant S. maltophilia strain D1585. Genome sequence analysis and functionality testing showed that DLP4 is a temperate phage capable of lysogenizing D1585. Two moron genes of interest, folA (BIT20_024) and ybiA (BIT20_065), were identified and investigated for their putative activities using complementation testing and phenotypic and transcriptomic changes between wild-type D1585 and the D1585::DLP4 lysogen. The gp24 / folA gene encodes dihydrofolate reductase (DHFR: FolA), an enzyme responsible for resistance to the antibiotic trimethoprim. I-TASSER analysis of DLP4 FolA predicted structural similarity to Bacillus anthracis DHFR and minimum inhibitory concentration experiments demonstrated that lysogenic conversion of D1585 by DLP4 provided the host cell with an increase in trimethoprim resistance. The gp65 / ybiA gene encodes N-glycosidase YbiA, which in E. coli BW25113 is required for its swarming motility phenotype. Expressing DLP4 ybiA in strain ybiA770(del)::kan restored its swarming motility activity to wildtype levels. Reverse transcription-PCR confirmed the expression of both of these genes during DLP4 lysogeny.ConclusionsS. maltophilia temperate phage DLP4 contributes to the antibiotic resistance exhibited by its lysogenized host strain. Genomic analyses can greatly assist in the identification of phage moron genes potentially involved in lysogenic conversion. Further research is required to fully understand the specific contributions temperate phage moron genes provide with respect to the antibiotic resistance and virulence of S. maltophilia host cells.

Dataset Information

Discovery of a New Microbial Origin Cold-Active Neopullulanase Capable for Effective Conversion of Pullulan to Panose.

Publications

Discovery of a New Microbial Origin Cold-Active Neopullulanase Capable for Effective Conversion of Pullulan to Panose.

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