Role of the cysteine protease interpain A of Prevotella intermedia in breakdown and release of haem from haemoglobin.

摘要: The gram-negative oral anaerobe Prevotella intermedia forms an iron(III) protoporphyrin IX pigment from haemoglobin. bacterium expresses a 90 kDa cysteine protease, InpA (interpain A), homologue of Streptococcus pyogenes streptopain (SpeB). role in haemoglobin breakdown and haem release was investigated. At pH 7.5, mediated oxidation oxyhaemoglobin to hydroxymethaemoglobin [in which the iron is oxidized Fe(III) state carries OH- as sixth co-ordinate ligand] by limited proteolysis globin chains indicated SDS/PAGE MALDI (matrix-assisted laser-desorption ionization)-TOF (time-of-flight) analysis. Prolonged incubation at 7.5 did not result further protein breakdown, but formation haemichrome (where Fe atom co-ordinated another amino acid ligand addition proximal histidine residue) resistant degradation InpA. InpA-mediated hydroxymethaemoglobin-agarose minimal compared with trypsin 7.5. 6.0, increased rate greater than auto-oxidation, producing aquomethaemoglobin (with water ligand), resulted its complete loss. Aquomethaemoglobin prevented blocking dissociation ligation azide, whereas haem-free rapid, even Both methaemoglobin were inhibited protease inhibitor E-64 [trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane]. In summary, we conclude that may play central acquisition mediating oxidation, degrading haem-globin affinity weakened under acidic conditions.