Poster: Heavy metals & phytoremediation

Abs # 43: Characterization of alr0975 from Nostoc sp. PCC7120, a prokaryotic gene similar to phytochelatin synthase

Presenter:	Tsuji, Naoki , ntsuji@phs.osaka-u.ac.jp
Authors	Tsuji, Naoki  (A)   Nishikori, Shingo  (B)   Iwabe, Osamu  (A)   Shiraki, Kentaro  (B)   Miyasaka, Hitoshi  (C)   Takagi, Masahiro  (B)   Hirata, Kazumasa  (A)   Miyamoto, Kazuhisa  (A)
Affiliations:	(A): Graduate School of Pharmaceutical Sciences, Osaka University (B): School of Materials Sciences, Japan Advanced Institute of Science and Technology (C): The Kansai Electric Power Co., Technical Research Center

Phytochelatins (PCs) are well known as the heavy metal-detoxifying peptides in eukaryotic higher plants, microalgae, fungi, and nematode. In contrast, neither PCs nor PC synthase genes have ever been identified in any prokaryotes including the cyanobacteria. The genome sequences for the cyanobacterium Nostoc sp. PCC 7120 were recently completed and allowed us to identify a gene encoding a PC synthase-like protein, termed alr0975. The predicted product of alr0975 contains the conserved N-terminal domain but not the variable C-terminal domain found in eukaryotic PC synthases. Despite the presence of this gene, no synthesis of PCs was observed in Nostoc cells challenged by Cd²⁺ or Zn²⁺, at any of the concentrations tested. Northern analysis revealed that alr0975 is not expressed under normal conditions nor is it induced in the presence of heavy metals. However, recombinant alr0975 protein strongly catalyzed the first step of PC synthesis, in which glutathione (GSH) is converted to γ-glutamylcysteine (γ-EC), although the protein only weakly catalyzed the second step of PC synthesis, namely the transfer of γ-EC moiety to an acceptor GSH molecule to form PC₂. Consistent with these results, Step 1 proceeded even in the absence of heavy metals in E. coli cells expressing alr0975, whereas synthesis of PCs and γ-EC required heavy metals in cells expressing the Arabidopsis PC synthase gene, AtPCS1, or a C-terminal deleted derivative, AtPCS1(Δ222-485), which retains only the N-terminal sequence homologous to alr0975. Based on these results, alr0975 appears to be a more primitive form of the PC synthases found in eukaryotes.