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Cite: NAR/gkaa742 2020



PUL General Information

Literature Information

PUL ID

PUL0017

PubMed

29093754, Biotechnol Biofuels. 2017 Oct 30;10:250. doi: 10.1186/s13068-017-0933-7. eCollection 2017.

Characterization method

qRT-PCR,Western Blot,isothermal titration calorimetry

Genomic accession number

NC_011898.1

Nucelotide position range

2466252-2481041

Substrate

cellobiose

Loci

CCEL_RS10665-CCEL_RS10705

Species

Ruminiclostridium cellulolyticum/1521

Degradation or Biosynthesis

degradation

Gene Name

Locus Tag

Protein ID

Gene Position

GenBank Contig Range

EC Number

tnpA CCEL_RS10665 WP_015926613.1 0 - 456 (-) NC_011898.1:2466252-2466708 -
- CCEL_RS10670 WP_015925554.1 717 - 3153 (-) NC_011898.1:2466969-2469405 -
- CCEL_RS10675 WP_015925555.1 3887 - 4715 (-) NC_011898.1:2470139-2470967 -
- CCEL_RS10680 WP_015925556.1 4701 - 5622 (-) NC_011898.1:2470953-2471874 -
- CCEL_RS10685 WP_015925557.1 5728 - 7078 (-) NC_011898.1:2471980-2473330 -
- CCEL_RS10690 WP_015925558.1 7413 - 9030 (-) NC_011898.1:2473665-2475282 -
- CCEL_RS10695 WP_015925559.1 9007 - 10891 (-) NC_011898.1:2475259-2477143 -
- CCEL_RS10700 WP_015925560.1 10945 - 12244 (-) NC_011898.1:2477197-2478496 -
- CCEL_RS10705 WP_015925561.1 12444 - 14790 (-) NC_011898.1:2478696-2481042 -

Cluster number

1

Gene name

Gene position

Gene type

Found by CGCFinder?

tnpA 1 - 456 (-) CDS No
- 718 - 3153 (-) CAZyme: GH94 Yes
- 3888 - 4715 (-) TC: gnl|TC-DB|P94530|3.A.1.1.34 Yes
- 4702 - 5622 (-) TC: gnl|TC-DB|P94529|3.A.1.1.34 Yes
- 5729 - 7078 (-) STP: STP|SBP_bac_1 Yes
- 7414 - 9030 (-) TF: DBD-Pfam|HTH_AraC,DBD-Pfam|HTH_AraC,DBD-SUPERFAMILY|0036286,DBD-SUPERFAMILY|0035607 Yes
- 9008 - 10891 (-) TC: gnl|TC-DB|F4LXP4|8.A.59.2.1 Yes
- 10946 - 12244 (-) STP: STP|SBP_bac_1 Yes
- 12445 - 14790 (-) TC: gnl|TC-DB|Q9FI56|3.A.9.1.2 Yes

PUL ID

PUL0017

PubMed

29093754, Biotechnol Biofuels. 2017 Oct 30;10:250. doi: 10.1186/s13068-017-0933-7. eCollection 2017.

Title

A seven-gene cluster in Ruminiclostridium cellulolyticum is essential for signalization, uptake and catabolism of the degradation products of cellulose hydrolysis.

Author

Fosses A, Mate M, Franche N, Liu N, Denis Y, Borne R, de Philip P, Fierobe HP, Perret S

Abstract

BACKGROUND: Like a number of anaerobic and cellulolytic Gram-positive bacteria, the model microorganism Ruminiclostridium cellulolyticum produces extracellular multi-enzymatic complexes called cellulosomes, which efficiently degrade the crystalline cellulose. Action of the complexes on cellulose releases cellobiose and longer cellodextrins but to date, little is known about the transport and utilization of the produced cellodextrins in the bacterium. A better understanding of the uptake systems and fermentation of sugars derived from cellulose could have a major impact in the field of biofuels production. RESULTS: We characterized a putative ABC transporter devoted to cellodextrins uptake, and a cellobiose phosphorylase (CbpA) in R. cellulolyticum. The genes encoding the components of the ABC transporter (a binding protein CuaA and two integral membrane proteins) and CbpA are expressed as a polycistronic transcriptional unit induced in the presence of cellobiose. Upstream, another polycistronic transcriptional unit encodes a two-component system (sensor and regulator), and a second binding protein CuaD, and is constitutively expressed. The products might form a three-component system inducing the expression of cuaABC and cbpA since we showed that CuaR is able to recognize the region upstream of cuaA. Biochemical analysis showed that CbpA is a strict cellobiose phosphorylase inactive on longer cellodextrins; CuaA binds to all cellodextrins (G2-G5) tested, whereas CuaD is specific to cellobiose and presents a higher affinity to this sugar. This results are in agreement with their function in transport and signalization, respectively. Characterization of a cuaD mutant, and its derivatives, indicated that the ABC transporter and CbpA are essential for growth on cellobiose and cellulose. CONCLUSIONS: For the first time in a Gram-positive strain, we identified a three-component system and a conjugated ABC transporter/cellobiose phosphorylase system which was shown to be essential for the growth of the model cellulolytic bacterium R. cellulolyticum on cellobiose and cellulose. This efficient and energy-saving system of transport and phosphorolysis appears to be the major cellobiose utilization pathway in R. cellulolyticum, and seems well adapted to cellulolytic life-style strain. It represents a new way to enable engineered strains to utilize cellodextrins for the production of biofuels or chemicals of interest from cellulose.