Structural highlights
FunctionPublication Abstract from PubMedHisA is a (betaalpha)8 barrel enzyme that catalyzes the Amadori rearrangement of ProFAR to PRFAR in the histidine biosynthesis pathway and it is a paradigm for the study of enzyme evolution. Still, its exact catalytic mechanism has remained unclear. Here, we present crystal structures of wild type Salmonella enterica HisA (SeHisA) in its apo state and of mutants D7N and D7N/D176A in complex with two different conformations of the labile substrate ProFAR, which was structurally visualized for the first time. Site-directed mutagenesis and kinetics demonstrated that Asp7 acts as the catalytic base and Asp176 as the catalytic acid. The SeHisA structures with ProFAR display two different states of the long loops on the catalytic face of the structure, and demonstrate that initial binding of ProFAR to the active site is independent of loop interactions. When the long loops enclose the substrate, ProFAR adopts an extended conformation where its non-reacting half is in a product-like conformation. This change is associated with shifts in a hydrogen-bond network including His47, Asp129, Thr171 and Ser202, all shown to be functionally important. The closed-conformation structure is highly similar to the bi-functional HisA homologue PriA in complex with PRFAR, thus proving that structure and mechanism are conserved between HisA and PriA. This study clarifies the mechanistic cycle of HisA and provides a striking example of how an enzyme and its substrate can undergo coordinated conformational changes before catalysis. Two-step Ligand Binding in a (betaalpha)8 Barrel Enzyme - Substrate-bound Structures Shed New Light on the Catalytic Cycle of HisA.,Soderholm A, Guo X, Newton MS, Evans GB, Nasvall J, Patrick WM, Selmer M J Biol Chem. 2015 Aug 20. pii: jbc.M115.678086. PMID:26294764[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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