Crystal structure of histidinol phosphate aminotransferase (HisC) from Escherichia coli, and its covalent complex with pyridoxal-5 '-phosphate and L-histidinol phosphate

The biosynthesis of histidine is a central metabolic process in organisms r anging from bacteria to yeast and plants. The seventh step in the synthesis of histidine within eubacteria is carried out by a pyridoxal-5'-phosphate (PLP)-dependent L-histidinol phosphate aminotransferase (HisC, EC 2.6.1.9). Here, we report the crystal structure of L-histidinol phosphate aminotrans ferase from Escherichia coli, as a complex with pyridoxamine-5'-phosphate ( PMP) at 1.5 Angstrom resolution, as the internal aldimine with PLP, and in a covalent, tetrahedral complex consisting of PLP and L-histidinol phosphat e attached to Lys214, both at 2.2 Angstrom resolution. This covalent comple x resembles, in structural terms, the gem-diamine intermediate that is form ed transiently during conversion of the internal to external aldimine. HisC is a dimeric enzyme with a mass of approximately 80 kDa. Like most PLP -dependent enzymes, each HisC monomer consists of two domains, a larger PLP -binding domain having an alpha/beta/alpha topology, and a smaller domain. An N-terminal arm contributes to the dimerization of the two monomers. The PLP-binding domain of HisC shows weak sequence similarity, but significant structural similarity with the PLP-binding domains of a number of PLP-depen dent enzymes. Residues that interact with the PLP cofactor, including Tyr55 , Asn157, Asp184, Tyr187, Ser213, Lys214 and Arg222, are conserved in the f amily of aspartate, tyrosine and histidinol phosphate aminotransferases. Th e imidazole ring Of L-histidinol phosphate is bound, in part, through a hyd rogen bond with Tyr110, a residue that is substituted by Phe in the broad s ubstrate specific HisC enzymes from Zymomonas mobilis and Bacillus subtilis . Comparison of the structures of the HisC internal aldimine, the PMP complex and the HisC L-histidinol phosphate complex reveal minimal changes in prot ein or ligand structure. Proton transfer, required for conversion of the ge m-diamine to the external aldimine, does not appear to be limited by the di stance between substrate and lysine amino groups. We propose that the tetra hedral complex has resulted from non-productive binding of L-histidinol pho sphate soaked into the HisC crystals, resulting in its inability to be conv erted to the external aldimine at the HisC active site. (C) 2001 Academic P ress.