Properties of guaiacol peroxidase activities isolated from corn root plasma membranes.
Although several investigations have demonstrated a plasma membrane (PM)-bound peroxidase activity in plants, this study is the first, to our knowledge, to purify and characterize the enzymes responsible. Proteins were extracted from highly enriched and thoroughly washed PMs. Washing and solubilization procedures indicated that the enzymes were tightly bound to the membrane. At least two distinct peroxidase activities could be separated by cation exchange chromatography (pmPOX1 and pmPOX2). Prosthetic groups were identified in fractions with peroxidase activity by absorption spectra, and the corresponding protein bands were identified by heme staining. The activities of the peroxidase enzymes responded different to various substrates and effectors and had different thermal stabilities and pH and temperature optima. Because the enzymes were localized at the PM and were not effected by p-chloromercuribenzoate, they were probably class III peroxidases. Additional size exclusion chromatography of pmPOX1 revealed a single activity peak with a molecular mass of 70 kD for the native enzyme, whereas pmPOX2 had two activity peaks (155 and 40 kD). Further analysis of these fractions by a modified sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with heme staining confirmed the estimated molecular masses of the size exclusion chromatography.
Mika
Angela
A
Universität Hamburg, Institut für Allgemeine Botanik, Ohnhorststrasse 18, D-22609 Hamburg, Germany.
Lüthje
Sabine
S
D002462:Cell Membrane / Q000201:enzymology*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D009195:Peroxidase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D018517:Plant Roots / Q000166:cytology* / Q000201:enzymology; D013379:Substrate Specificity; D013696:Temperature; D003313:Zea mays / Q000166:cytology* / Q000201:enzymology*
C121048:guaiacol peroxidase; D009195:Peroxidase