title: Metabolic engineering to expand the substrate spectrum of Pseudomonas putida toward sucrose.
abstract: Sucrose is an important disaccharide used as a substrate in many industrial applications. It is a major component of molasses, a cheap by-product of the sugar industry. Unfortunately, not all industrially relevant organisms, among them Pseudomonas putida, are capable of metabolizing sucrose. We chose a metabolic engineering approach to circumvent this blockage and equip P. putida with the activities necessary to consume sucrose. Therefore, we constructed a pair of broad-host range mini-transposons (pSST - sucrose splitting transposon), carrying either cscA, encoding an invertase able to split sucrose into glucose and fructose, or additionally cscB, encoding a sucrose permease. Introduction of cscA was sufficient to convey sucrose consumption and the additional presence of cscB had no further effect, though the sucrose permease was built and localized to the membrane. Sucrose was split extracellularly by the activity of the invertase CscA leaking out of the cell. The transposons were also used to confer sucrose consumption to Cupriavidus necator. Interestingly, in this strain, CscB acted as a glucose transporter, such that C. necator also gained the ability to grow on glucose. Thus, the pSST transposons are functional tools to extend the substrate spectrum of Gram-negative bacterial strains toward sucrose.
authors.affiliation: Fachgebiet für Systembiotechnologie, Fakultät für Maschienenwesen, Technische Universität München, Garching, Germany.
authors.forename: Hannes
authors.initials: H
authors.lastname: Löwe
authors.affiliation: Fachgebiet für Systembiotechnologie, Fakultät für Maschienenwesen, Technische Universität München, Garching, Germany.
authors.forename: Lukas
authors.initials: L
authors.lastname: Schmauder
authors.affiliation: Fachgebiet für Systembiotechnologie, Fakultät für Maschienenwesen, Technische Universität München, Garching, Germany.
authors.forename: Karina
authors.initials: K
authors.lastname: Hobmeier
authors.affiliation: Fachgebiet für Systembiotechnologie, Fakultät für Maschienenwesen, Technische Universität München, Garching, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Kremling
authors.affiliation: Fachgebiet für Systembiotechnologie, Fakultät für Maschienenwesen, Technische Universität München, Garching, Germany.
authors.forename: Katharina
authors.identifier: 0000-0002-8334-353X
authors.initials: K
authors.lastname: Pflüger-Grau
mesh_terms: D003470:Culture Media / Q000737:chemistry; D020118:Cupriavidus necator / Q000235:genetics / Q000378:metabolism; D004251:DNA Transposable Elements; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D060847:Metabolic Engineering; D016254:Mutagenesis, Insertional; D016958:Pseudomonas putida / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013395:Sucrose / Q000378:metabolism*; D013549:Sweetening Agents / Q000378:metabolism*; D043324:beta-Fructofuranosidase / Q000235:genetics / Q000378:metabolism
chemical_list: D003470:Culture Media; D004251:DNA Transposable Elements; D026901:Membrane Transport Proteins; D011994:Recombinant Proteins; D013549:Sweetening Agents; D013395:Sucrose; D043324:beta-Fructofuranosidase
keywords: 
  Pseudomonas putida
  ; metabolic engineering; sucrose metabolism