Microbial production and chemical transformation of poly-γ-glutamate.
Bottom Line: The occurrence of multiple carboxyl residues in PGA likely plays a role in its relative unsuitability for the development of bio-nylon plastics and thus, establishment of an efficient PGA-reforming strategy is of great importance.Aside from the potential applications of PGA proposed to date, a new technique for chemical transformation of PGA is also discussed.Finally, some techniques for PGA and its derivatives in advanced material technology are presented.
Affiliation: Agricultural Science, Graduate School of Integrated Arts and Sciences, Kochi University, Nankoku, Kochi, 783-8502, Japan.Show MeSH
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Mentions: Elucidation of the mechanism reproducible for the synthesis of PGA would be indispensable for obtaining a better understanding of enzymes involved in its synthesis. Based on the structural features of PGA, such as the introduction of non-proteinaceous d-glutamate and its unusual γ-amide linkage formula, the existence of a novel enzyme that can catalyze non-ribosomal glutamate ligation (viz., polymerization) is predicted. Moreover, the nucleotide formed by coincident ATP hydrolysis is ADP, not AMP (Ashiuchi et al., 2001b; Urushibata et al., 2002), revealing that PGA is synthesized in an amide-ligation manner (Ashiuchi, 2010). The amide-ligation mechanism is catalyzed by typical amide ligases with a Rossmann-like fold such as murein-biosynthetic enzymes (Eveland et al., 1997), or ATP-dependent (ADP-forming) carboxylate-amine/thiol ligases (peptide synthetases) with ATP-grasp domain(s), including glutathione synthetase and d-alanine-d-alanine ligase (Galperin and Koonin, 1997). Both types of amide ligases are commonly characterized by a lack of isomerization activity for amino acid residues in a growing chain, resulting in the substrate having the same stereochemistry as the polymer produced. Therefore, d-amino acid residues in polyamides generated via the amide-ligation mechanism will be derived from free d-amino acids in cells (Ashiuchi et al., 2013b). Ashiuchi and colleagues (2004) actually found a membrane-associated DL-PGA synthetic activity from B. subtilis subsp. chungkookjang, in which both d- and l-glutamate served as direct substrates. Interestingly, there is a noteworthy difference in the proposed catalytic mechanisms of amide ligases (Fig. 2), namely that the Rossmann-type enzymes activate the C-terminal carboxyl residue of the polymers (as the acceptor in peptide elongation; Sheng et al., 2000), whereas the ATP-grasp-type enzymes generally phosphorylate the carboxyl group of donor substrates (Fan et al., 1995; Kino et al., 2009). This may sometimes cause a lack of stereo-exactitude in the former enzymes, resulting in dl-copolymer production. Ashiuchi and colleagues (2001b) previously observed that there was no phosphorylation activity for the monomers of glutamate during the elongation reaction with a B. subtilis DL-PGA synthetase, predicting that the enzyme will belong to the superfamily of Rossmann-type amide ligases (Eveland et al., 1997).
Affiliation: Agricultural Science, Graduate School of Integrated Arts and Sciences, Kochi University, Nankoku, Kochi, 783-8502, Japan.