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. 1999 Jul;65(7):3071-4.
doi: 10.1128/AEM.65.7.3071-3074.1999.

Purification and characterization of a secreted laccase of Gaeumannomyces graminis var. tritici

W A Edens  1 , T Q Goins, D Dooley, J M Henson
Affiliations

Purification and characterization of a secreted laccase of Gaeumannomyces graminis var. tritici

W A Edens et al. Appl Environ Microbiol. 1999 Jul.

Abstract

We purified a secreted fungal laccase from filtrates of Gaeumannomyces graminis var. tritici cultures induced with copper and xylidine. The active protein had an apparent molecular mass of 190 kDa and yielded subunits with molecular masses of 60 kDa when denatured and deglycosylated. This laccase had a pI of 5.6 and an optimal pH of 4.5 with 2,6-dimethoxyphenol as its substrate. Like other, previously purified laccases, this one contained several copper atoms in each subunit, as determined by inductively coupled plasma spectroscopy. The active enzyme catalyzed the oxidation of 2, 6-dimethoxyphenol (Km = 2.6 x 10(-5) +/- 7 x 10(-6) M), catechol (Km = 2.5 x 10(-4) +/- 1 x 10(-5) M), pyrogallol (Km = 3.1 x 10(-4) +/- 4 x 10(-5) M), and guaiacol (Km = 5.1 x 10(-4) +/- 2 x 10(-5) M). In addition, the laccase catalyzed the polymerization of 1, 8-dihydroxynaphthalene, a natural fungal melanin precursor, into a high-molecular-weight melanin and catalyzed the oxidation, or decolorization, of the dye poly B-411, a lignin-like polymer. These findings indicate that this laccase may be involved in melanin polymerization in this phytopathogen's hyphae and/or in lignin depolymerization in its infected plant host.

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Figures

FIG. 1
FIG. 1
Determination of molecular masses of purified G. graminis var. tritici laccase and laccase subunits. (A) Lane 1, molecular size standards (silver stain); lane 2, native laccase (silver stain); lane 3, native laccase (activity stain with DMOP); lane 4, native and denatured laccase (silver stain). (B) Lane 1, molecular size standards (silver stain); lane 2, denatured laccase (silver stain); lane 3, denatured, deglycosylated laccase (60 kDa) and PNGase F deglycosylase (36 kDa; silver stain).
FIG. 2
FIG. 2
Decolorization of poly B-411 dye by laccase. Decolorization was monitored at different times by observing the absorbance ratios at 593 and 483 nm. Squares, control reaction (no laccase added); diamonds, reactions where laccase was added. Results shown are averages.
FIG. 3
FIG. 3
Polymerization of 1,8-DHN by laccase, monitored by scanning changes in absorbance from 320 to 520 nm.
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References

    1. Bell A, Wheeler M. Biosynthesis and functions of fungal melanins. Annu Rev Phytopathol. 1986;24:411–451.
    1. Blum H, Beier H, Gross H. Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis. 1987;8:93–99.
    1. Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72:248–254. - PubMed
    1. Chet I, Trojanowski J, Huttermann A. Decolourisation of the dye poly B-411 and its correlation with lignin degradation by fungi. Microbios Lett. 1985;29:37–43.
    1. Claus H, Filip Z. The evidence of a laccase-like enzyme activity in a Bacillus sphaericus strain. Microbiol Res. 1997;152:209–216.

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