title: Poly-alpha-glutamic acid synthesis using a novel catalytic activity of RimK from Escherichia coli K-12.
abstract: Poly-L-α-amino acids have various applications because of their biodegradable properties and biocompatibility. Microorganisms contain several enzymes that catalyze the polymerization of L-amino acids in an ATP-dependent manner, but the products from these reactions contain amide linkages at the side residues of amino acids: e.g., poly-γ-glutamic acid, poly-ε-lysine, and cyanophycin. In this study, we found a novel catalytic activity of RimK, a ribosomal protein S6-modifying enzyme derived from Escherichia coli K-12. This enzyme catalyzed poly-α-glutamic acid synthesis from unprotected L-glutamic acid (Glu) by hydrolyzing ATP to ADP and phosphate. RimK synthesized poly-α-glutamic acid of various lengths; matrix-assisted laser desorption ionization-time of flight-mass spectrometry showed that a 46-mer of Glu (maximum length) was synthesized at pH 9. Interestingly, the lengths of polymers changed with changing pH. RimK also exhibited 86% activity after incubation at 55°C for 15 min, thus showing thermal stability. Furthermore, peptide elongation seemed to be catalyzed at the C terminus in a stepwise manner. Although RimK showed strict substrate specificity toward Glu, it also used, to a small extent, other amino acids as C-terminal substrates and synthesized heteropeptides. In addition, RimK-catalyzed modification of ribosomal protein S6 was confirmed. The number of Glu residues added to the protein varied with pH and was largest at pH 9.5.
authors.affiliation: Department of Applied Chemistry, Faculty of Science and Engineering, Waseda University, 3-4-1 Ohkubo, Shinjuku-ku, Tokyo 169-8555, Japan. kkino@waseda.jp
authors.forename: Kuniki
authors.initials: K
authors.lastname: Kino
authors.forename: Toshinobu
authors.initials: T
authors.lastname: Arai
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Arimura
mesh_terms: D002384:Catalysis; D048168:Escherichia coli K12 / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D018698:Glutamic Acid / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000737:chemistry*; D011107:Polylysine / Q000138:chemical synthesis* / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D029968:Escherichia coli Proteins; D010455:Peptides; D011107:Polylysine; D018698:Glutamic Acid