Abstract
The second and third steps of the histidine biosynthetic pathway (HBP) in plants are catalyzed by a bifunctional enzyme–HISN2. The enzyme consists of two distinct domains, active respectively as a phosphoribosyl-AMP cyclohydrolase (PRA-CH) and phosphoribosyl-ATP pyrophosphatase (PRA-PH). The domains are analogous to single-domain enzymes encoded by bacterial hisI and hisE genes, respectively. The calculated sequence similarity networks between HISN2 analogs from prokaryotes and eukaryotes suggest that the plant enzymes are closest relatives of those in the class of Deltaproteobacteria. In this work, we obtained crystal structures of HISN2 enzyme from Medicago truncatula (MtHISN2) and described its architecture and interactions with AMP. The AMP molecule bound to the PRA-PH domain shows positioning of the N1-phosphoribosyl relevant to catalysis. AMP bound to the PRA-CH domain mimics a part of the substrate, giving insights into the reaction mechanism. The latter interaction also arises as a possible second-tier regulatory mechanism of the HBP flux, as indicated by inhibition assays and isothermal titration calorimetry.
Highlights
The second and third steps of the histidine biosynthetic pathway (HBP) in plants are catalyzed by a bifunctional enzyme–HISN2
Studies of the HBP helped to discover the regulation of amino acid biosynthesis by a ttenuation[10]
Fungal trifunctional proteins (HIS4 in yeast) with phosphoribosyl-ATP pyrophosphatase (PRA-PH), phosphoribosyl-AMP cyclohydrolase (PRA-CH), and HDH activities derive from orders close to Myxococcales
Summary
The second and third steps of the histidine biosynthetic pathway (HBP) in plants are catalyzed by a bifunctional enzyme–HISN2. The HBP is rather conservative among different domains of life; there are differences in the number of genes involved in the pathway and their expression p attern[11] In bacteria, his genes are arranged in a compact operon (hisGDC [NB] HAF [IE]), with three of them (hisD, hisNB and hisIE) sometimes but not always coding for bifunctional e nzymes[10,12]. Recent progress in molecular biology techniques has revealed that many of the enzymatic steps of the HBP in plants are performed by proteins encoded by single genes, which is in contrast to the extensive gene redundancy found in other amino acid biosynthetic pathways in plants[14]. Five of the HBP enzymes in A. thaliana are encoded by single-copy genes, with duplications in HISN1, HISN5, and HISN616
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