title: Calcium-dependent heme structure in the reduced forms of the bacterial cytochrome c peroxidase from Paracoccus pantotrophus.
abstract: This work reports for the first time a resonance Raman study of the mixed-valence and fully reduced forms of Paracoccus pantotrophus bacterial cytochrome c peroxidase. The spectra of the active mixed-valence enzyme show changes in the structure of the ferric peroxidatic heme compared to the fully oxidized enzyme; these differences are observed upon reduction of the electron-transferring heme and upon full occupancy of the calcium site. For the mixed-valence form in the absence of Ca(2+), the peroxidatic heme is six-coordinate and low-spin on the basis of the frequencies of the structure-sensitive Raman lines: the enzyme is inactive. With added Ca(2+), the peroxidatic heme is five-coordinate high-spin and active. The calcium-dependent spectral differences indicate little change in the conformation of the ferrous electron-transferring heme, but substantial changes in the conformation of the ferric peroxidatic heme. Structural changes associated with Ca(2+) binding are indicated by spectral differences in the structure-sensitive marker lines, the out-of-plane low-frequency macrocyclic modes, and the vibrations associated with the heme substituents of that heme. The Ca(2+)-dependent appearance of a strong gamma 15 saddling-symmetry mode for the mixed-valence form is consistent with a strong saddling deformation in the active peroxidatic heme, a feature seen in the Raman spectra of other peroxidases. For the fully reduced form in the presence of Ca(2+), the resonance Raman spectra show that the peroxidatic heme remains high-spin.
authors.affiliation: REQUIMTE/CQFB, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica, Portugal.
authors.forename: Sofia R
authors.initials: SR
authors.lastname: Pauleta
authors.forename: Yi
authors.initials: Y
authors.lastname: Lu
authors.forename: Celia F
authors.initials: CF
authors.lastname: Goodhew
authors.forename: Isabel
authors.initials: I
authors.lastname: Moura
authors.forename: Graham W
authors.initials: GW
authors.lastname: Pettigrew
authors.forename: John A
authors.initials: JA
authors.lastname: Shelnutt
mesh_terms: D001671:Biochemistry / Q000379:methods; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002412:Cations; D003578:Cytochrome-c Peroxidase / Q000378:metabolism*; D004789:Enzyme Activation; D006418:Heme / Q000737:chemistry*; D007477:Ions; D008274:Magnesium / Q000737:chemistry; D008956:Models, Chemical; D008968:Molecular Conformation; D010100:Oxygen / Q000737:chemistry; D042084:Paracoccus pantotrophus / Q000201:enzymology*; D011487:Protein Conformation; D013059:Spectrum Analysis, Raman / Q000379:methods
chemical_list: D002412:Cations; D007477:Ions; D006418:Heme; D003578:Cytochrome-c Peroxidase; D008274:Magnesium; D010100:Oxygen; D002118:Calcium