Publications

• Structural, kinetic, and evolutionary peculiarities of HISN3, a plant 5'-ProFAR isomerase.

  Witek W., Imiolczyk B., Ruszkowski M.

  Histidine biosynthesis is essential for the growth and development of plants, where it occurs within chloroplasts. The eleven reactions are catalyzed by eight enzymes, known as HISN1-8, each acting sequentially. Here, we present the crystal structures of a 5'-ProFAR isomerase (HISN3) from the mode ... >> More

  Histidine biosynthesis is essential for the growth and development of plants, where it occurs within chloroplasts. The eleven reactions are catalyzed by eight enzymes, known as HISN1-8, each acting sequentially. Here, we present the crystal structures of a 5'-ProFAR isomerase (HISN3) from the model legume Medicago truncatula bound to its enzymatically synthesized substrate (ProFAR) and product (PrFAR). The active site of MtHISN3 contains a sodium cation that participates in ligand recognition, a feature not observed in bacterial and fungal structures of homologous enzymes. The steady-state kinetics of wild-type MtHISN3 revealed a slightly higher turnover rate compared to its bacterial homologs. Plant HISN3 sequences contain an unusually elongated Lys60-Ser91 fragment, while deletion of the 74-80 region resulted in a 30-fold loss in catalytic efficiency compared to the wild-type. Molecular dynamics simulations suggested that the fragment facilitates product release, thereby contributing to a higher k_cat. Moreover, conservation analyses suggested a non-cyanobacterial origin for plant HISN3 enzymes, which is another instance of a non-cyanobacterial enzyme in the plant histidine biosynthetic pathway. Finally, a virtual screening campaign yielded five molecules, with the energy gains ranging between -13.6 and -13.1 kcal/mol, which provide new scaffolds for the future development of herbicides. << Less

  Plant Physiol Biochem 215:109065-109065(2024) [PubMed] [EuropePMC]

• Crystal structure of the yeast His6 enzyme suggests a reaction mechanism.

  Quevillon-Cheruel S., Leulliot N., Graille M., Blondeau K., Janin J., van Tilbeurgh H.

  The Saccharomyces cerevisiae His6 gene codes for the enzyme phosphoribosyl-5-amino-1-phosphoribosyl-4-imidazolecarboxamide isomerase, catalyzing the fourth step in histidine biosynthesis. To get an insight into the structure and function of this enzyme, we determined its X-ray structure at a resol ... >> More

  The Saccharomyces cerevisiae His6 gene codes for the enzyme phosphoribosyl-5-amino-1-phosphoribosyl-4-imidazolecarboxamide isomerase, catalyzing the fourth step in histidine biosynthesis. To get an insight into the structure and function of this enzyme, we determined its X-ray structure at a resolution of 1.30 A using the anomalous diffraction signal of the protein's sulphur atoms at 1.77 A wavelength. His6 folds in an (alpha/beta)8 barrel similar to HisA, which performs the same function in bacteria and archaea. We found a citrate molecule from the buffer bound in a pocket near the expected position of the active site and used it to model the open form of the substrate (phosphoribulosyl moiety), which is a reaction intermediate. This model enables us to identify catalytic residues and to propose a reaction mechanism where two aspartates act as acid/base catalysts: Asp134 as a proton donor for ring opening, and Asp9 as a proton acceptor and donor during enolization of the aminoaldose. Asp9 is conserved in yeast His6 and bacterial or archaeal HisA sequences, and Asp134 has equivalents in both HisA and TrpF, but they occur at a different position in the protein sequence. << Less

  Protein Sci. 15:1516-1521(2006) [PubMed] [EuropePMC]

• Two (betaalpha)(8)-barrel enzymes of histidine and tryptophan biosynthesis have similar reaction mechanisms and common strategies for protecting their labile substrates.

  Henn-Sax M., Thoma R., Schmidt S., Hennig M., Kirschner K., Sterner R.

  The enzymes N'-[(5'-phosphoribosyl)formimino]-5-aminoimidazole-4-carboxamide ribonucleotide isomerase (HisA) and phosphoribosylanthranilate isomerase (TrpF) are sugar isomerases that are involved in histidine and tryptophan biosynthesis, respectively. Both enzymes have the (betaalpha)(8)-barrel fo ... >> More

  The enzymes N'-[(5'-phosphoribosyl)formimino]-5-aminoimidazole-4-carboxamide ribonucleotide isomerase (HisA) and phosphoribosylanthranilate isomerase (TrpF) are sugar isomerases that are involved in histidine and tryptophan biosynthesis, respectively. Both enzymes have the (betaalpha)(8)-barrel fold and catalyze Amadori rearrangements of a thermolabile aminoaldose into the corresponding aminoketose. To identify those amino acids that are essential for catalysis, conserved residues at the active sites of both HisA and TrpF from the hyperthermophile Thermotoga maritima were replaced by site-directed mutagenesis, and the purified variants were investigated by steady-state enzyme kinetics. Aspartate 8, aspartate 127, and threonine 164 appeared to be important for the HisA reaction, whereas cysteine 7 and aspartate 126 appeared to be important for the TrpF reaction. On the basis of these results and the X-ray structure of a complex between TrpF and a bound product analogue, a reaction mechanism involving general acid-base catalysis and a Schiff base intermediate is proposed for both enzymes. A comparison of the HisA and TrpF enzymes from T. maritima and Escherichia coli showed that, at the physiological temperatures of 80 and 37 degrees C, respectively, the enzymes from the hyperthermophile have significantly higher catalytic efficiencies than the corresponding enzymes from mesophiles. These results suggest that HisA and TrpF have similar chemical reaction mechanisms and use the same strategy to prevent the loss of their thermolabile substrates. << Less

  Biochemistry 41:12032-12042(2002) [PubMed] [EuropePMC]