Construction and analysis of a recombinant cyanobacterium expressing a chromosomally inserted gene for an ethylene-forming enzyme at the psbAI locus.

The coding sequence of a gene for a Pseudomonas syringae ethylene-forming enzyme was inserted at the psbAI locus in a cyanobacterium, Synechococcus elongatus PCC 7942 via rps12-mediated gene replacement. The recombinant strain photoautotrophically produced ethylene at 451 nl ml(-1) h(-1) OD730(-1), but showed a depressed specific growth rate as well as a yellow-green phenotype indicating a severe metabolic stress. The rate of ethylene production in the recombinant culture decreased as a result of competition with faster growing ethylene-non-forming mutants that carried short nucleotide insertions within the coding sequence of the gene for the ethylene-forming enzyme.

[1]  R. Haselkorn,et al.  Expression of a family of psbA genes encoding a photosystem II polypeptide in the cyanobacterium Anacystis nidulans R2. , 1986, The EMBO journal.

[2]  K. Takahama,et al.  Gene replacement in cyanobacteria mediated by a dominant streptomycin-sensitive rps12 gene that allows selection of mutants free from drug resistance markers. , 2001, Microbiology.

[3]  S. Tanase,et al.  Molecular cloning in Escherichia coli, expression, and nucleotide sequence of the gene for the ethylene-forming enzyme of Pseudomonas syringae pv. phaseolicola PK2. , 1992, Biochemical and biophysical research communications.

[4]  K. Nagahama,et al.  Heterologous expression of the gene for the ethylene-forming enzyme fromPseudomonas syringae in the cyanobacteriumSynechococcus , 2005, Biotechnology Letters.

[5]  A. van der Ende,et al.  A host-vector system for gene cloning in the cyanobacterium Anacystis nidulans R2. , 1983, Plasmid.

[6]  S. Golden,et al.  Functional Elements of the Strong psbAIPromoter of Synechococcus elongatus PCC 7942 , 2001, Journal of bacteriology.

[7]  R. Rippka Isolation and purification of cyanobacteria. , 1988, Methods in enzymology.

[8]  M. Matsuoka,et al.  Photosynthetic conversion of carbon dioxide to ethylene by the recombinant cyanobacterium, Synechococcus sp. PCC 7942, which harbors a gene for the ethylene-forming enzyme of Pseudomonas syringae , 1997 .

[9]  Wang,et al.  A method of graphically analyzing substrate-inhibition kinetics. , 1999, Biotechnology and bioengineering.

[10]  S. Tanase,et al.  Two reactions are simultaneously catalyzed by a single enzyme: the arginine-dependent simultaneous formation of two products, ethylene and succinate, from 2-oxoglutarate by an enzyme from Pseudomonas syringae. , 1992, Biochemical and biophysical research communications.

[11]  P. Weisbeek,et al.  Identification of replication and stability functions in the complete nucleotide sequence of plasmid pUH24 from the cyanobacterium Synechococcus sp. PCC 7942 , 1992, Molecular microbiology.

[12]  S. Tanase,et al.  Purification and properties of an ethylene-forming enzyme from Pseudomonas syringae pv. phaseolicola PK2. , 1991, Journal of general microbiology.

[13]  B. Grimm,et al.  Structural genes of glutamate 1-semialdehyde aminotransferase for porphyrin synthesis in a cyanobacterium and Escherichia coli , 2004, Molecular and General Genetics MGG.