Characterization of a low redox potential laccase from the basidiomycete C30.
A new exocellular laccase was purified from the basidiomycete C30. LAC2 is an acidic protein (pI = 3.2) preferentially produced upon a combined induction by copper and p-hydroxybenzoate. The spectroscopic signature (UV/visible and EPR) of this isoform is typical of multicopper oxidases, but its enzymatic and physico-chemical properties proved to be markedly different from those of LAC1, the constitutive laccase previously purified from the same organism. In particular, the LAC2 kcat values observed for the oxidation of the substrates syringaldazine (kcat = 65 600 min-1), ABTS (2,2-azino-bis-[3-ethylthiazoline-6-sulfonate] (kcat = 41 000 min-1) and guaiacol (kcat = 75 680 min-1) are 10-40 times those obtained with LAC1 and the redox potential of its T1 copper is 0.17 V lower than that of LAC1 (E degrees = 0.73 V). This is the first report on a single organism producing simultaneously both a high and a low redox potential laccase. The cDNA, clac2, was cloned and sequenced. It encodes a protein of 528 amino acids that shares 69% identity (79% similarity) with LAC1 and 81% identity (95% similarity) with Lcc3-2 from Polyporus ciliatus (AF176321-1), its nearest neighbor in database. Possible reasons for why this basidiomycete produces, in vivo, enzyme forms with such different behaviors are discussed.
Klonowska
Agnieszka
A
Laboratoire de Bioinorganique Structurale CNRS UMR 6517 and Laboratoire d'Ecologie Microbienne, CNRS UMR 6116, Faculté des Sciences de St Jérôme, Marseille, France.
Gaudin
Christian
C
Fournel
André
A
Asso
Marcel
M
Le Petit
Jean
J
Giorgi
Michel
M
Tron
Thierry
T
D001386:Azides / Q000494:pharmacology; D001487:Basidiomycota / Q000201:enzymology*; D018076:DNA, Complementary / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D042845:Laccase; D008958:Models, Molecular; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D013053:Spectrophotometry; D013329:Structure-Activity Relationship; D014466:Ultraviolet Rays
D001386:Azides; D018076:DNA, Complementary; D010455:Peptides; D020033:Protein Isoforms; D012333:RNA, Messenger; D010088:Oxidoreductases; D042845:Laccase