Metabolic engineering to expand the substrate spectrum of Pseudomonas putida toward sucrose.
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.
['Löwe|Hannes|H|', 'Schmauder|Lukas|L|', 'Hobmeier|Karina|K|', 'Kremling|Andreas|A|', 'Pflüger-Grau|Katharina|K|0000-0002-8334-353X']
D003470:Culture Media D020118:Cupriavidus necator D004251:DNA Transposable Elements D026901:Membrane Transport Proteins D060847:Metabolic Engineering D016254:Mutagenesis, Insertional D016958:Pseudomonas putida D011994:Recombinant Proteins D013395:Sucrose D013549:Sweetening Agents D043324:beta-Fructofuranosidase
metabolic engineering Pseudomonas putida sucrose metabolism