title: Construction of a chassis for hydrogen production: physiological and molecular characterization of a Synechocystis sp. PCC 6803 mutant lacking a functional bidirectional hydrogenase.
abstract: Cyanobacteria are photosynthetic prokaryotes that are promising 'low-cost' microbial cell factories due to their simple nutritional requirements and metabolic plasticity, and the availability of tools for their genetic manipulation. The unicellular non-nitrogen-fixing Synechocystis sp. PCC 6803 is the best studied cyanobacterial strain and its genome was the first to be sequenced. The vast amount of physiological and molecular data available, together with a relatively small genome, makes Synechocystis suitable for computational metabolic modelling and to be used as a photoautotrophic chassis in synthetic biology applications. To prepare it for the introduction of a synthetic hydrogen producing device, a Synechocystis sp. PCC 6803 deletion mutant lacking an active bidirectional hydrogenase (ΔhoxYH) was produced and characterized at different levels: physiological, proteomic and transcriptional. The results showed that, under conditions favouring hydrogenase activity, 17 of the 210 identified proteins had significant differential fold changes in comparisons of the mutant with the wild-type. Most of these proteins are related to the redox and energy state of the cell. Transcriptional studies revealed that only six genes encoding those proteins exhibited significant differences in transcript levels. Moreover, the mutant exhibits similar growth behaviour compared with the wild-type, reflecting Synechocystis plasticity and metabolic adaptability. Overall, this study reveals that the Synechocystis ΔhoxYH mutant is robust and can be used as a photoautotrophic chassis for the integration of synthetic constructs, i.e. molecular constructs assembled from well characterized biological and/or synthetic parts (e.g. promoters, regulators, coding regions, terminators) designed for a specific purpose.
authors.affiliation: Faculdade de Ciências, Departamento de Biologia, Universidade do Porto, Rua do Campo Alegre, Edifício FC4, 4169-007 Porto, Portugal.|IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal.
authors.forename: Filipe
authors.initials: F
authors.lastname: Pinto
authors.affiliation: Biological and Environmental Systems Group, ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Mapping Street, Sheffield S1 3JD, UK.
authors.forename: Karin A
authors.initials: KA
authors.lastname: van Elburg
authors.affiliation: IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal.
authors.forename: Catarina C
authors.initials: CC
authors.lastname: Pacheco
authors.affiliation: IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal.
authors.forename: Miguel
authors.initials: M
authors.lastname: Lopo
authors.affiliation: Biological and Environmental Systems Group, ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Mapping Street, Sheffield S1 3JD, UK.
authors.forename: Josselin
authors.initials: J
authors.lastname: Noirel
authors.affiliation: Instituto Universitario de Matemática Pura y Aplicada, Universidad Politécnica de Valencia, Camí de Vera, E-46071 Valencia, Spain.
authors.forename: Arnau
authors.initials: A
authors.lastname: Montagud
authors.affiliation: Instituto Universitario de Matemática Pura y Aplicada, Universidad Politécnica de Valencia, Camí de Vera, E-46071 Valencia, Spain.
authors.forename: Javier F
authors.initials: JF
authors.lastname: Urchueguía
authors.affiliation: Biological and Environmental Systems Group, ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Mapping Street, Sheffield S1 3JD, UK.
authors.forename: Phillip C
authors.initials: PC
authors.lastname: Wright
authors.affiliation: Faculdade de Ciências, Departamento de Biologia, Universidade do Porto, Rua do Campo Alegre, Edifício FC4, 4169-007 Porto, Portugal.|IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal.
authors.forename: Paula
authors.initials: P
authors.lastname: Tamagnini
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006859:Hydrogen / Q000378:metabolism*; D006864:Hydrogenase / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D046939:Synechocystis / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D006859:Hydrogen; D006864:Hydrogenase