Purification and spectroscopic studies on catechol oxidase from lemon balm (Melissa officinalis).
A catechol oxidase from lemon balm (Melissa officinalis) moCO which only catalyzes the oxidation of catechols to quinones without hydroxylating tyrosine was purified. The molecular mass of the M. officinalis enzyme of 39,370 Da was obtained by MALDI mass spectrometry and the isoelectric point was determined to be 3.4. Addition of 2 eq. H(2)O(2) to the enzyme leads to oxy catechol oxidase. In the UV/Vis spectrum two new absorption bands occur at 343 nm (ε=8510 M(-1)cm(-1)) and 580 nm (ε=580 M(-1)cm(-1)) due to O(2)(2-)Cu (II) charge transfer transitions in accordance with the oxy forms of other type 3 copper proteins. The N-terminal sequence has been determined by Edman degradation to NPVQAPELDKCGTAT, exhibiting a proline at the second and sixth position conserved in other polyphenol oxidases.
Institut für Biophysikalische Chemie, Universität Wien, Wien, Austria. annette.rompel@univie.ac.at
Annette
A
Rompel
Klaudia
K
Büldt-Karentzopoulos
Christian
C
Molitor
Bernt
B
Krebs
D000595:Amino Acid Sequence; D020134:Catalytic Domain; D004156:Catechol Oxidase / Q000737:chemistry / Q000302:isolation & purification*; D003300:Copper / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006861:Hydrogen Peroxide / Q000737:chemistry; D007526:Isoelectric Point; D031334:Melissa / Q000737:chemistry / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity
D010940:Plant Proteins; D003300:Copper; D006861:Hydrogen Peroxide; D004156:Catechol Oxidase