scholarly journals CphA2 is a novel type of cyanophycin synthetase in N2-fixing cyanobacteria

Microbiology ◽  
2016 ◽  
Vol 162 (3) ◽  
pp. 526-536 ◽  
Author(s):  
Friederike Klemke ◽  
Dennis J. Nürnberg ◽  
Karl Ziegler ◽  
Gabriele Beyer ◽  
Uwe Kahmann ◽  
...  
Keyword(s):  
Cyanophycin Synthetase

scholarly journals Incorporation of alternative amino acids into cyanophycin by different cyanophycin synthetases heterologously expressed in Corynebacterium glutamicum

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ramona Wördemann ◽  
Lars Wiefel ◽  
Volker F. Wendisch ◽  
Alexander Steinbüchel
Keyword(s):  
Amino Acids ◽  
Dry Weight ◽  
Growth Conditions ◽  
Amino Acid Biosynthesis ◽  
Water Soluble ◽  
Lysine Content ◽  
Polyaspartic Acid ◽  
Acid Biosynthesis ◽  
Cyanophycin Synthetase ◽  
Potential Precursor

AbstractCyanophycin (multi-l-arginyl-poly-l-aspartic acid; also known as cyanophycin grana peptide [CGP]) is a biopolymer that could be used in various fields, for example, as a potential precursor for the synthesis of polyaspartic acid or for the production of CGP-derived dipeptides. To extend the applications of this polymer, it is therefore of interest to synthesize CGP with different compositions. A recent re-evaluation of the CGP synthesis in C. glutamicum has shown that C. glutamicum is a potentially interesting microorganism for CGP synthesis with a high content of alternative amino acids. This study shows that the amount of alternative amino acids can be increased by using mutants of C. glutamicum with altered amino acid biosynthesis. With the DM1729 mutant, the lysine content in the polymer could be increased up to 33.5 mol%. Furthermore, an ornithine content of up to 12.6 mol% was achieved with ORN2(Pgdh4). How much water-soluble or insoluble CGP is synthesized is strongly related to the used cyanophycin synthetase. CphADh synthesizes soluble CGP exclusively. However, soluble CGP could also be isolated from cells expressing CphA6308Δ1 or CphA6308Δ1_C595S in addition to insoluble CGP in all examined strains. The point mutation in CphA6308Δ1_C595S partially resulted in a higher lysine content. In addition, the CGP content could be increased to 36% of the cell dry weight under optimizing growth conditions in C. glutamicum ATCC13032. All known alternative major amino acids for CGP synthesis (lysine, ornithine, citrulline, and glutamic acid) could be incorporated into CGP in C. glutamicum.

scholarly journals A mutant of the cyanobacteriumAnabaena variabilisATCC 29413 lacking cyanophycin synthetase: growth properties and ultrastructural aspects

2001 ◽  
Vol 196 (1) ◽  
pp. 13-18 ◽  
Author(s):  
Karl Ziegler ◽  
Dirk P. Stephan ◽  
Elfriede K. Pistorius ◽  
Hans G. Ruppel ◽  
Wolfgang Lockau
Keyword(s):  
Growth Properties ◽  
Cyanophycin Synthetase

Molecular characterization of the cyanophycin synthetase from Synechocystis sp. strain PCC6308

2000 ◽  
Vol 174 (5) ◽  
pp. 297-306 ◽  
Author(s):  
Elsayed Aboulmagd ◽  
Fred Bernd Oppermann-Sanio ◽  
Alexander Steinbüchel
Keyword(s):  
Molecular Characterization ◽  
Cyanophycin Synthetase ◽  
Synechocystis Sp

Partial purification and characterization of a non-cyanobacterial cyanophycin synthetase from Acinetobacter calcoaceticus strain ADP1 with regard to substrate specificity, substrate affinity and binding to cyanophycin

Microbiology ◽  
2004 ◽  
Vol 150 (8) ◽  
pp. 2599-2608 ◽  
Author(s):  
Martin Krehenbrink ◽  
Alexander Steinbüchel
Keyword(s):  
Aspartic Acid ◽  
Biochemical Characterization ◽  
Acinetobacter Calcoaceticus ◽  
Partial Purification ◽  
Atp Binding ◽  
Substrate Affinity ◽  
Two Dimensional Gel Electrophoresis ◽  
Atp Binding Site ◽  
Cyanophycin Synthetase

This study reports, for the first time, purification and biochemical characterization of a cyanophycin synthetase from a non-cyanobacterial strain. Cyanophycin synthetase of Acinetobacter calcoaceticus strain ADP1 was purified 69-fold from recombinant Escherichia coli by two chromatographic steps and one novel affinity step utilizing the Mg2+-dependent binding of the enzyme to cyanophycin. Unlike cyanobacterial cyanophycin synthetases characterized so far, the purified enzyme from A. calcoaceticus strain ADP1 did not accept lysine as an alternative substrate to arginine. The apparent K m-values for arginine (47 μM) and aspartic acid (240 μM) were similar to those of known cyanophycin synthetases from cyanobacteria, but this enzyme had a slightly higher affinity for aspartic acid. In addition, the two different ATP-binding sites of the enzyme were characterized independently of each other with respect to K m values for ATP. The ATP-binding site responsible for the addition of arginine was found to have a much higher affinity for ATP (38 μM) than that responsible for the addition of aspartate (210 mM). Furthermore, the binding of the enzyme to the two possible forms of cyanophycin granule polypeptide (CGP), CGP-Asp and CGP-Arg, was studied. While both forms bound around 30–40 % of the enzyme activity present under the assay conditions, binding was Mg2+-dependent in the case of CGP-Asp. Two-dimensional gel electrophoresis revealed that both forms of cyanophycin were equally abundant in cyanophycin-accumulating cells of A. calcoaceticus ADP1.

scholarly journals Production of cyanophycin in Rhizopus oryzae through the expression of a cyanophycin synthetase encoding gene

2011 ◽  
Vol 93 (3) ◽  
pp. 1167-1174 ◽  
Author(s):  
Bas J. Meussen ◽  
Ruud A. Weusthuis ◽  
Johan P. M. Sanders ◽  
Leo H. de Graaff
Keyword(s):  
Rhizopus Oryzae ◽  
Encoding Gene ◽  
Cyanophycin Synthetase

scholarly journals Effect of Nitrogen Source on Cyanophycin Synthesis in Synechocystis sp. Strain PCC 6308

2006 ◽  
Vol 188 (3) ◽  
pp. 934-940 ◽  
Author(s):  
Nancy H. Kolodny ◽  
Deborah Bauer ◽  
Kerstyn Bryce ◽  
Kristen Klucevsek ◽  
Amy Lane ◽  
...  
Keyword(s):  
Nitrogen Source ◽  
Nitrogen Limitation ◽  
Nitrogen Starvation ◽  
Cellular Protein ◽  
Proton Nuclear Magnetic Resonance ◽  
Resonance Spectroscopy ◽  
Nitrogen Incorporation ◽  
Cyanophycin Synthetase ◽  
Synechocystis Sp

ABSTRACT Experiments were carried out to examine the effects of nitrogen source on nitrogen incorporation into cyanophycin during nitrogen limitation and repletion, both with or without inhibition of protein synthesis, in cyanobacteria grown on either nitrate or ammonium. The use of nitrate and ammonium, 14N labeled in the growth medium and 15N labeled in the repletion medium, allows the determination of the source of nitrogen in cyanophycin using proton nuclear magnetic resonance spectroscopy. The data suggest that nitrogen from both the breakdown of cellular protein (14N) and directly from the medium (15N) is incorporated into cyanophycin. Nitrogen is incorporated into cyanophycin at different rates and to different extents, depending on the source of nitrogen (ammonium or nitrate) and whether the cells are first starved for nitrogen. These differences appear to be related to the activity of nitrate reductase in cells and to the possible expression of cyanophycin synthetase during nitrogen starvation.

scholarly journals Metabolic Engineering of Saccharomyces cerevisiae for Production of Novel Cyanophycins with an Extended Range of Constituent Amino Acids

2009 ◽  
Vol 75 (11) ◽  
pp. 3437-3446 ◽  
Author(s):  
Anna Steinle ◽  
Klaus Bergander ◽  
Alexander Steinbüchel
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Ornithine Carbamoyltransferase ◽  
Argininosuccinate Synthetase ◽  
Argininosuccinate Lyase ◽  
Arginine Metabolism ◽  
Extended Range ◽  
Putative Precursor ◽  
Cyanophycin Synthetase

ABSTRACT Cyanophycin (multi-l-arginyl-poly-l-aspartic acid; also known as cyanophycin grana peptide [CGP]) is a putative precursor for numerous biodegradable technically used chemicals. Therefore, the biosynthesis and production of the polymer in recombinant organisms is of special interest. The synthesis of cyanophycin derivatives consisting of a wider range of constituents would broaden the applications of this polymer. We applied recombinant Saccharomyces cerevisiae strains defective in arginine metabolism and expressing the cyanophycin synthetase of Synechocystis sp. strain PCC 6308 in order to synthesize CGP with citrulline and ornithine as constituents. Strains defective in arginine degradation (Car1 and Car2) accumulated up to 4% (wt/wt) CGP, whereas strains defective in arginine synthesis (Arg1, Arg3, and Arg4) accumulated up to 15.3% (wt/wt) of CGP, which is more than twofold higher than the previously content reported in yeast and the highest content ever reported in eukaryotes. Characterization of the isolated polymers by different analytical methods indicated that CGP synthesized by strain Arg1 (with argininosuccinate synthetase deleted) consisted of up to 20 mol% of citrulline, whereas CGP from strain Arg3 (with ornithine carbamoyltransferase deleted) consisted of up to 8 mol% of ornithine, and CGP isolated from strain Arg4 (with argininosuccinate lyase deleted) consisted of up to 16 mol% lysine. Cultivation experiments indicated that the incorporation of citrulline or ornithine is enhanced by the addition of low amounts of arginine (2 mM) and also by the addition of ornithine or citrulline (10 to 40 mM), respectively, to the medium.

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