STRUCTURE OF BARLEY-GRAIN PEROXIDASE REFINED AT 1.9-ANGSTROM RESOLUTION - A PLANT PEROXIDASE REVERSIBLY INACTIVATED AT NEUTRAL PH

The crystal structure of the major peroxidase of barley grain (BP 1) h as been solved by molecular replacement and phase combination and refi ned to an R-factor of 19.2% for all data between 38 and 1.9 Angstrom, The refined model includes amino acid residues 1-309, one calcium ion, one sodium ion, iron-protoporphyrin IX, and 146 solvent molecules, BP 1 has the apparently unique property of being unable to catalyze the reaction with the primary substrate hydrogen peroxide to form compound I at pH values > 5, a feature investigated by obtaining crystal struc ture data at pH 5.5, 7.5, and 8.5, Structural comparison shows that th e overall fold of inactive barley grain peroxidase at these pH values resembles that of both horseradish peroxidase C and peanut peroxidase, The key differences between the structures of active horseradish pero xidase C and inactive BP 1 include the orientation of the catalytic di stal histidine, disruption of a hydrogen bond between this histidine a nd a conserved asparagine, and apparent substitution of calcium at the distal cation binding site with sodium at pH 7.5. These profound chan ges are a result of a dramatic structural rearrangement to the loop re gion between helices B and C, This is the first time that structural r earrangements linked to active site chemistry have been observed by cr ystallography in the peroxidase domain distal to heme.