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Cite: NAR/gkaa742 2020



PUL General Information

Literature Information

PUL ID

PUL0024

PubMed

31915220, mSphere. 2020 Jan 8;5(1):e00771-19. doi: 10.1128/mSphere.00771-19.

Characterization method

enzyme activity assay,qPCR,substrate binding assay

Genomic accession number

HE717008.1

Nucelotide position range

1-14712

Substrate

kestose

Loci

CCG34774.1-CCG34784.1

Species

uncultured bacterium/77133

Degradation or Biosynthesis

degradation

Gene Name

Locus Tag

Protein ID

Gene Position

GenBank Contig Range

EC Number

- - CCG34773.1 0 - 107 (+) HE717008.1:1-108 -
- - CCG34774.1 568 - 1423 (+) HE717008.1:569-1424 -
- - CCG34775.1 1565 - 2996 (-) HE717008.1:1566-2997 -
- - CCG34776.1 3041 - 4973 (-) HE717008.1:3042-4974 -
- - CCG34777.1 5175 - 6225 (-) HE717008.1:5176-6226 -
- - CCG34778.1 6369 - 8076 (-) HE717008.1:6370-8077 -
- - CCG34779.1 8075 - 9956 (-) HE717008.1:8076-9957 -
- - CCG34780.1 10112 - 10955 (+) HE717008.1:10113-10956 -
- - CCG34781.1 11356 - 11944 (-) HE717008.1:11357-11945 -
- - CCG34782.1 11945 - 12173 (-) HE717008.1:11946-12174 -
- - CCG34783.1 12455 - 13859 (-) HE717008.1:12456-13860 -
- - CCG34784.1 14001 - 14712 (-) HE717008.1:14002-14713 -

Cluster number

1

Gene name

Gene position

Gene type

Found by CGCFinder?

- 1 - 107 (+) CDS No
- 569 - 1423 (+) CDS No
- 1566 - 2996 (-) CAZyme: GH32 Yes
- 3042 - 4973 (-) TC: gnl|TC-DB|Q8NMD6|4.A.1.2.12 Yes
- 5176 - 6225 (-) TF: DBD-Pfam|LacI,DBD-SUPERFAMILY|0036955 Yes
- 6370 - 8076 (-) TC: gnl|TC-DB|Q9WYC4|3.A.1.135.5 Yes
- 8076 - 9956 (-) TC: gnl|TC-DB|Q9WYC3|3.A.1.135.5 Yes
- 10113 - 10955 (+) TF: DBD-Pfam|HTH_1,DBD-SUPERFAMILY|0040266 Yes
- 11357 - 11944 (-) other Yes
- 11946 - 12173 (-) TF: DBD-Pfam|HTH_3,DBD-SUPERFAMILY|0043143 Yes
- 12456 - 13859 (-) TC: gnl|TC-DB|P50487|2.A.40.2.1 Yes
- 14002 - 14712 (-) CDS No

PUL ID

PUL0024

PubMed

31915220, mSphere. 2020 Jan 8;5(1):e00771-19. doi: 10.1128/mSphere.00771-19.

Title

Harvesting of Prebiotic Fructooligosaccharides by Nonbeneficial Human Gut Bacteria.

Author

Wang Z, Tauzin AS, Laville E, Tedesco P, Letisse F, Terrapon N, Lepercq P, Mercade M, Potocki-Veronese G

Abstract

Prebiotic oligosaccharides, such as fructooligosaccharides, are increasingly being used to modulate the composition and activity of the gut microbiota. However, carbohydrate utilization analyses and metagenomic studies recently revealed the ability of deleterious and uncultured human gut bacterial species to metabolize these functional foods. Moreover, because of the difficulties of functionally profiling transmembrane proteins, only a few prebiotic transporters have been biochemically characterized to date, while carbohydrate binding and transport are the first and thus crucial steps in their metabolization. Here, we describe the molecular mechanism of a phosphotransferase system, highlighted as a dietary and pathology biomarker in the human gut microbiome. This transporter is encoded by a metagenomic locus that is highly conserved in several human gut Firmicutes, including Dorea species. We developed a generic strategy to deeply analyze, in vitro and in cellulo, the specificity and functionality of recombinant transporters in Escherichia coli, combining carbohydrate utilization locus and host genome engineering and quantification of the binding, transport, and growth rates with analysis of phosphorylated carbohydrates by mass spectrometry. We demonstrated that the Dorea fructooligosaccharide transporter is specific for kestose, whether for binding, transport, or phosphorylation. This constitutes the biochemical proof of effective phosphorylation of glycosides with a degree of polymerization of more than 2, extending the known functional diversity of phosphotransferase systems. Based on these new findings, we revisited the classification of these carbohydrate transporters.IMPORTANCE Prebiotics are increasingly used as food supplements, especially in infant formulas, to modify the functioning and composition of the microbiota. However, little is currently known about the mechanisms of prebiotic recognition and transport by gut bacteria, while these steps are crucial in their metabolism. In this study, we established a new strategy to profile the specificity of oligosaccharide transporters, combining microbiomics, genetic locus and strain engineering, and state-of-the art metabolomics. We revisited the transporter classification database and proposed a new way to classify these membrane proteins based on their structural and mechanistic similarities. Based on these developments, we identified and characterized, at the molecular level, a fructooligosaccharide transporting phosphotransferase system, which constitutes a biomarker of diet and gut pathology. The deciphering of this prebiotic metabolization mechanism by a nonbeneficial bacterium highlights the controversial use of prebiotics, especially in the context of chronic gut diseases.