Molecular characterization of cyanophycin synthetase, the enzyme catalyzing the biosynthesis of the cyanobacterial reserve material multi-L-arginyl-poly-L-aspartate (cyanophycin).
Cyanophycin (multi-L-arginyl-poly-L-aspartate), a water-insoluble reserve polymer of cyanobacteria, is a product of nonribosomal peptide synthesis. The purification of cyanophycin synthetase of the cyanobacterium Anabaena variabilis is described. In sodium dodecylsulfate/polyacrylamide gel electrophoresis, the enzyme preparation shows one band with an apparent molecular mass of 100 kDa. The native enzyme has an apparent molecular mass of approximately 230 kDa, as determined by size-exclusion chromatography, suggesting that the active form is a homodimer. During catalysis, ATP is converted to ADP. The gene coding for cyanophycin synthetase has been identified in the sequenced genome of Synechocystis sp. PCC 6803. The C-terminal 60% of the deduced amino acid sequence of cyanophycin synthetase show sequence similarity to enzymes of the superfamily of ligases involved in the biosynthesis of murein and of folyl-poly(gamma-glutamate). Cells of Escherichia coli harbouring the gene on a plasmid express active synthetase and accumulate cyanophycin-like material. The results prove that a single enzyme catalyzes the de novo synthesis of cyanophycin.
Ziegler
K
K
Biochemie der Pflanzen, Humboldt-Universität, Berlin, Germany.
Diener
A
A
Herpin
C
C
Richter
R
R
Deutzmann
R
R
Lockau
W
W
D000255:Adenosine Triphosphate / Q000378:metabolism; D017033:Anabaena / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry*; D003001:Cloning, Molecular; D000458:Cyanobacteria / Q000201:enzymology; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D010453:Peptide Synthases / Q000737:chemistry*; D010940:Plant Proteins / Q000096:biosynthesis*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D010940:Plant Proteins; D011994:Recombinant Proteins; C024250:cyanophycin; D000255:Adenosine Triphosphate; D010453:Peptide Synthases; C113257:cyanophycin synthase, bacteria