Structure and expression of a laccase gene from the ligninolytic basidiomycete Ceriporiopsis subvermispora

Karahanian, Eduardo; Corsini, Gino; Lobos, Sergio; Vicuña, Rafael

doi:10.1016/s0167-4781(98)00197-3

Cited by 68 publications

(39 citation statements)

References 36 publications

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“…Nine multicopper (MCO)-encoding C. subvermispora genes may be relevant to lignin degradation. Multiple alignments emphasizing signature regions (26,27) revealed the presence of seven laccases, in the strictest sense, one of which was previously known (28). This observation is in distinct contrast to the P. chrysosporium genome, which contains no laccases (12) (Fig.…”

Section: Resultsmentioning

confidence: 85%

Comparative genomics of Ceriporiopsis subvermispora and Phanerochaete chrysosporium provide insight into selective ligninolysis

Fernández‐Fueyo

Ruíz-Dueñas

Ferreira

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Efficient lignin depolymerization is unique to the wood decay basidiomycetes, collectively referred to as white rot fungi. Phanerochaete chrysosporium simultaneously degrades lignin and cellulose, whereas the closely related species, Ceriporiopsis subvermispora, also depolymerizes lignin but may do so with relatively little cellulose degradation. To investigate the basis for selective ligninolysis, we conducted comparative genome analysis of C. subvermispora and P. chrysosporium . Genes encoding manganese peroxidase numbered 13 and five in C. subvermispora and P. chrysosporium , respectively. In addition, the C. subvermispora genome contains at least seven genes predicted to encode laccases, whereas the P. chrysosporium genome contains none. We also observed expansion of the number of C. subvermispora desaturase-encoding genes putatively involved in lipid metabolism. Microarray-based transcriptome analysis showed substantial up-regulation of several desaturase and MnP genes in wood-containing medium. MS identified MnP proteins in C. subvermispora culture filtrates, but none in P. chrysosporium cultures. These results support the importance of MnP and a lignin degradation mechanism whereby cleavage of the dominant nonphenolic structures is mediated by lipid peroxidation products. Two C. subvermispora genes were predicted to encode peroxidases structurally similar to P. chrysosporium lignin peroxidase and, following heterologous expression in Escherichia coli , the enzymes were shown to oxidize high redox potential substrates, but not Mn 2+ . Apart from oxidative lignin degradation, we also examined cellulolytic and hemicellulolytic systems in both fungi. In summary, the C. subvermispora genetic inventory and expression patterns exhibit increased oxidoreductase potential and diminished cellulolytic capability relative to P. chrysosporium .

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Section: Resultsmentioning

confidence: 85%

Comparative genomics of Ceriporiopsis subvermispora and Phanerochaete chrysosporium provide insight into selective ligninolysis

Fernández‐Fueyo

Ruíz-Dueñas

Ferreira

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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265

235

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“…RT-PCR was used to isolate a fulllength cDNA as described by Karahanian et al (1998). Design of primers was based on the genomic sequence of lcs-1 (GenBank accession number AF053472).…”

Section: Methodsmentioning

confidence: 99%

“…Analytical IEF and protein concentration measurements were as described by Larrondo et al (2001); RNA extraction and Northern blot hybridization were as described by Karahanian et al (1998). The DNA probe corresponding to the entire coding region of Lcs-1 was labelled with [a-32 P]dCTP by nick translation (Gibco-BRL).…”

Section: Methodsmentioning

confidence: 99%

Heterologous expression of laccase cDNA from Ceriporiopsis subvermispora yields copper-activated apoprotein and complex isoform patterns

et al. 2003

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Analysis of genomic clones encoding a putative laccase in homokaryon strains of Ceriporiopsis subvermispora led to the identification of an allelic variant of the previously described lcs-1 gene. A cDNA clone corresponding to this gene was expressed in Aspergillus nidulans and in Aspergillus niger. Enzyme assays and Western blots showed that both hosts secreted active laccase. Relative to the isozymic forms of the native C. subvermispora enzyme, the A. niger-produced laccase had a higher molecular mass and gave a single band on IEF gels. In contrast, A. nidulans transformants secreted several isoforms remarkably similar to those of the native system. Considered together with previously reported Southern blots and protein sequencing, expression in A. nidulans supports the view that C. subvermispora has a single laccase gene and that multiple isoforms result from post-translational processes. In addition, several lines of evidence strongly suggest that under copper limitation, A. nidulans secretes apoprotein which can be reconstituted by a short incubation with Cu(I) and to a lesser extent with Cu(II).

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“…In addition, laccase production can be influenced by the nitrogen concentration in the culture medium (Gianfreda et al, 1999) and by the carbon source employed (Galhaup et al, 2002). Recently, an important effect of copper on laccase synthesis in Trametes versicolor, Pleurotus ostreatus and Ceriporiopsis subvermispora has been described (Collins & Dobson, 1997 ;Karahanian et al, 1998 ;Palmieri et al, 2000). In these studies it was shown that the addition of copper to the growth medium significantly increases laccase formation and that the regulation of the synthesis of several different laccase isoforms by copper occurs at the level of gene transcription.…”

Section: Abbreviationsmentioning

confidence: 99%

Characterization of the major laccase isoenzyme from Trametes pubescens and regulation of its synthesis by metal ions a aThe GenBank accession numbers for the lap2 and lap1a genes reported in this paper are AF414807 and AF414808, respectively.

et al. 2002

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The major laccase isoenzyme LAP2 secreted by the white-rot basidiomycete Trametes pubescens in response to high copper concentrations was purified to apparent electrophoretic homogeneity using anion-exchange chromatography and gel filtration. The monomeric protein has a molecular mass of 65 kDa, of which 18 % is glycosylation, and a pI value of 2 6. The pH optima of the laccase depend on the substrates oxidized and show bell-shaped pH activity profiles with an optimum of 3-4 5 for phenolic substrates such as 2,6-dimethoxyphenol or syringaldazine, while the non-phenolic substrates ABTS [2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid)] and ferrocyanide show a monotonic pH profile with a rate increasing with decreasing pH. The catalytic efficiencies k cat /K m determined for some of its substrates were 48W10 6 , 47W10 6 , 20W10 6 and 7W10 6 M N1 s N1 for ABTS, syringaldazine, ferrocyanide and oxygen, respectively. Furthermore, the gene lap2 encoding the purified laccase was cloned and its nucleotide sequence determined. The gene consists of 1997 bp, with the coding sequence interrupted by eight introns and flanked by an upstream region in which putative CAAT, TATA, MRE and CreA consensus sequences were identified. Based on Northern analysis containing total RNA from both induced and uninduced cultures, expression of lap2 is highly induced by copper, which is also corroborated by an increase in laccase activity in response to copper. A stimulating effect of various other heavy metal ions on laccase synthesis was also observed. In addition to induction, a second regulatory mechanism seems to be repression of lap2 transcription by glucose.

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Structure and expression of a laccase gene from the ligninolytic basidiomycete Ceriporiopsis subvermispora

Cited by 68 publications

References 36 publications

Comparative genomics of Ceriporiopsis subvermispora and Phanerochaete chrysosporium provide insight into selective ligninolysis

Comparative genomics of Ceriporiopsis subvermispora and Phanerochaete chrysosporium provide insight into selective ligninolysis

Heterologous expression of laccase cDNA from Ceriporiopsis subvermispora yields copper-activated apoprotein and complex isoform patterns

Characterization of the major laccase isoenzyme from Trametes pubescens and regulation of its synthesis by metal ions a aThe GenBank accession numbers for the lap2 and lap1a genes reported in this paper are AF414807 and AF414808, respectively.

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