Identification of the Anabaena sp.Strain PCC7120 Cyanophycin Synthetase as Suitable Enzyme for Production of Cyanophycin in Gram-Negative Bacteria Like Pseudomonas putida and Ralstonia eutropha

摘要

The Cyanophycin synthetase gene cphAl encoding the major Cyanophycin synthetase(CphA)of Anabaena sp.strain PCC7120 was expressed in Escherichia coli conferring so far the highest specific CphA activity to E.coli(6.7 nmol arginine per min and mg protein).CphAl and cphA genes of Synechocystis sp.strains PCC6803 and PCC6308 and Synechococcus strain MA 19 were also expressed in wild types and polyhydroxyalkanoate-negative(PHA)mutants of Pseudomonas putida and Ralstonia eutropha.Recombinant strains of these bacteria expressing cphAl accumulated generally more Cyanophycin(23.0 and 20.0% of cellular dry matter,CDM,respectively)than recombinants expressing any other cphA(6.8,9.0,or 15.8% of CDM for P.putida strains and 7.3,12.6,or 14.1% of CDM for R.eutropha).Furthermore,PHA-negative mutants of P.putida(9.7,10.0,17.5,or 24.0% of CDM)and R.eutropha(8.9,13.8,16.0,or 22.0% of CDM)accumulated generally more Cyanophycin than the corresponding PHA-positive parent strains(6.8,9.0,15.8,and 23.0% of CDM for P.putida strains and 7.3,12.6,14.1,or 20.0% of CDM for R.eutropha strains).Recombinant strains of Gram-positive bacteria(Bacillus megaterium,Corynebact erium glutamicum)were not suitable for Cyanophycin production due to accumulation of less Cyanophycin and retarded release of cyanophycin.PHA-negative mutants of P.putida and R.eutropha expressing cphAl of Anabaena sp.strain PCC7120 are therefore preferred candidates for industrial production of cyanophycin.