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abstracts
The Escherichia coli 6-carboxytetrahydropterin synthase (eCTPS), a homolog of 6-pyruvoyl tetrahydropterin synthase (PTPS), possesses a much stronger catalytic activity to cleave the side chain of sepiapterin in vitro rather than the genuine PTPS activity and catalyzes the conversion of dihydroneopterin triphosphate to 6-carboxy-5,6,7,8-tetrahydropterin in vivo. We have determined crystal structures of a wild type apo-eCTPS and a Cys27Ala mutant eCTPS complexed with sepiapterin up to 2.3 and 2.5 Å, respectively. The structures are highly conserved at the active site and the Zn2+ binding site. However, comparison of the eCTPS structures with those of mammalian PTPS homologs revealed that two specific residues Trp51 and Phe55, not existing in the mammalian PTPS, kept the substrate bound by stacking it with their side chains. Replacements of these two residues by site-directed mutagenesis to the residues, Met and Leu, existing only in mammalian PTPS, converted the eCTPS to have the mammalian PTPS activity. Our studies confirm that these two aromatic residues in eCTPS play an essential role in stabilizing the substrate and for the specific enzyme activity different from the original PTPS activity. These aromatic residues Trp51 and Phe55 are a key signature of bacterial PTPS enzymes that distinguish them from mammalian PTPS homologs.
abstracts
Glyoxylate cycle is a branched metabolic pathway in the TCA cycle that was initially discovered in microorganisms. The branched cycle plays an essential role in those organisms by providing the means for microorganisms to utilize acetate, ethanol, or fatty acids as carbon sources. In fact, pathogenic microorganisms rely on the glyoxylate cycle, rather than the TCA cycle, during infection. Therefore, the enzymes in the glyoxylate cycle of pathogens were suggested to be one of drug target molecules. Magnaporthe grisea isocitrate lyase (MgICL), a key enzyme in the cycle, is highly expressed during appressorium-mediated plant infection. In order to characterize the structural and functional features of MgICL, a structure of MgICL was determined at 2.7 Å resolution by X-ray crystallography. Recently, we are carrying out structure determination of MgICL in complex with a possible candidate for inhibitors. Our study could provide detailed structural features of MgICL and the binding mode of an inhibitor. This work was supported by a grant from Center for Fungal Pathogenesis by National Research Foundation, Republic of Korea.
