scholarly journals Localised laccase activity modulates distribution of lignin polymers in gymnosperm compression wood

2021 ◽  
Author(s):  
Hideto Hiraide ◽  
Yuki Tobimatsu ◽  
Arata Yoshinaga ◽  
Pui Ying Lam ◽  
Masaru Kobayashi ◽  
...  
Keyword(s):  
Compression Wood ◽  
Laccase Activity

In situ detection of laccase activity and immunolocalisation of a compression-wood-specific laccase (CoLac1) in differentiating xylem of Chamaecyparis obtusa

2016 ◽  
Vol 43 (6) ◽  
pp. 542 ◽  
Author(s):  
Hideto Hiraide ◽  
Masato Yoshida ◽  
Saori Sato ◽  
Hiroyuki Yamamoto
Keyword(s):  
Cell Wall ◽  
Secondary Wall ◽  
Compression Wood ◽  
Laccase Activity ◽  
Outer Part ◽  
Chamaecyparis Obtusa ◽  
Wall Layer ◽  
Wall Thickening ◽  
Lignin Concentration

The secondary cell wall of compression wood tracheids has a highly lignified region (S2 L) in its outermost portion. To better understand the mechanism of S2 L formation, we focussed on the activity of laccase (a monolignol oxidase) and performed in situ studies of this enzyme in differentiating compression wood. Staining of differentiating compression wood demonstrated that laccase activity began in all cell wall layers before the onset of lignification. We detected no activity of peroxidase (another monolignol oxidase) in any cell wall layer. Thus, laccase likely plays the major role in monolignol oxidisation during compression wood differentiation. Laccase activity was higher in the S2 L region than in other secondary wall regions, suggesting that this enzyme was responsible for the high lignin concentration in this region of the cell wall. Immunolabelling demonstrated the expression of a compression-wood-specific laccase (CoLac1) immediately following the onset of secondary wall thickening, this enzyme was localised to the S2 L region, whereas much less abundant in the S1 layer or inner S2 layer. Thus, the CoLac1 protein is most likely localised to the outer part of S2 and responsible for the high lignin concentration in the S2 L region.

scholarly journals Cerrena unicolor Laccases, Genes Expression and Regulation of Activity

Biomolecules ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 468
Author(s):  
Anna Pawlik ◽  
Beata Ciołek ◽  
Justyna Sulej ◽  
Andrzej Mazur ◽  
Przemysław Grela ◽  
...  
Keyword(s):  
Laccase Activity ◽  
Environmental Changes ◽  
Specific Activity ◽  
Electrochemical Analysis ◽  
White Rot ◽  
White Rot Fungus ◽  
Cerrena Unicolor ◽  
Genes Expression ◽  
Regulation Of Activity ◽  
Laccase Genes

A white rot fungus Cerrena unicolor has been identified as an important source of laccase, unfortunately regulation of this enzyme genes expression is poorly understood. Using 1D and 2D PAGE and LC-MS/MS, laccase isoenzymes were investigated in the liquid filtrate of C. unicolor culture. The level of expression of laccase genes was measured using qPCR. The elevated concentrations of copper and manganese in the medium caused greatest change in genes expression and three laccase transcripts were significantly affected after culture temperature was decreased from 28 to 4 °C or increased to 40 °C. The small differences in the PAGE band intensities of individual laccase proteins were also observed, indicating that given compound affect particular laccase’s transcript. Analyses of laccase-specific activity, at all tested conditions, showed the increased activities as compared to the control, suggesting that enzyme is regulated at the post-translational stage. We observed that the aspartic protease purified from C. unicolor, significantly stimulate laccase activity. Moreover, electrochemical analysis of protease-treated laccase sample had 5 times higher redox peaks. The obtained results indicate that laccases released by C. unicolor are regulated at transcriptional, translational, and at the post-translational steps of gene expression helping fungus adapt to the environmental changes.

scholarly journals Comprehensive analysis of the isozyme composition of laccase derived from Japanese lacquer tree, Toxicodendron vernicifluum

2021 ◽  
Vol 67 (1) ◽  
Author(s):  
Mariko Takano ◽  
Masaya Nakamura ◽  
Masanobu Tabata
Keyword(s):  
Isoelectric Focusing ◽  
Laccase Activity ◽  
Maximum Activity ◽  
Acetone Powder ◽  
Blue Color ◽  
Buffer Solution ◽  
Activity Staining ◽  
Water Extracts ◽  
Ph Range ◽  
Toxicodendron Vernicifluum

AbstractWe performed an analysis using isoelectric focusing to comprehensively clarify the isozyme composition of laccase derived from Japanese lacquer tree, Toxicodendron vernicifluum. When water extracts of acetone powder obtained from lacquer were subjected to isoelectric focusing, five bands within pI 7.35–9.30 and nine bands within pI 3.50–5.25 were detected using Coomassie staining. Similarly, laccase activity staining using guaiacol showed five bands within pI 7.35–9.30 and three bands within pI 3.50–4.25. However, laccase activity staining using gallic acid showed remarkable staining within pI 3.50–5.85, whereas staining was very weak within pI 7.35–9.30. When the water extracts of acetone powder were fractionated into the fractions containing bands within pI 7.35–9.30 and pI 3.50–5.85 by SP-Sepharose column chromatography, the former had a blue color and the latter a yellow color. The laccase activity was measured for each of the fractions in buffer solution in the pH range of 2.5–8.0. When syringaldazine, guaiacol, and 2,6-dimethoxyphenol were used as substrates, the yellow fraction showed considerably higher activity than the blue fraction for pH 5.5–7.5. When 3-methylcatechol and 4-methylcatechol were used as substrates, the yellow fraction showed higher activity for pH 4.5–6.5, and the blue fraction showed higher activity for pH 7.0–8.0. When 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) was used as the substrate, both fractions showed maximum activity at optimum pH of 3.0–4.0. Conventionally, in research on blue laccase derived from lacquer, the non-blue fraction corresponding to the yellow fraction lower than pI 6 has been removed during the purification process and thus has not been analyzed. Our results indicated that yellow laccase was present in the non-blue components of lacquer and that it may play a role in urushiol polymerization with previously reported blue laccase.

NUMERICAL AND EXPERIMENTAL STUDIES ON INFLUENCE OF COMPRESSION WOOD TIMBER DISTORTION

2000 ◽  
Vol 18 (8) ◽  
pp. 1897-1919 ◽  
Author(s):  
S. Ormarssonand ◽  
H. Petersson ◽  
O. Dahlblom
Keyword(s):  
Compression Wood ◽  
Experimental Studies

scholarly journals Protein Engineering Approaches to Enhance Fungal Laccase Production in S. cerevisiae

2021 ◽  
Vol 22 (3) ◽  
pp. 1157
Author(s):  
Pablo Aza ◽  
Felipe de Salas ◽  
Gonzalo Molpeceres ◽  
David Rodríguez-Escribano ◽  
Iñigo de la Fuente ◽  
...  
Keyword(s):  
Protein Engineering ◽  
Directed Evolution ◽  
Signal Peptide ◽  
Laccase Activity ◽  
Laccase Production ◽  
Wide Range ◽  
Mating Factor ◽  
Conserved Amino Acids ◽  
Basidiomycete Fungi

Laccases secreted by saprotrophic basidiomycete fungi are versatile biocatalysts able to oxidize a wide range of aromatic compounds using oxygen as the sole requirement. Saccharomyces cerevisiae is a preferred host for engineering fungal laccases. To assist the difficult secretion of active enzymes by yeast, the native signal peptide is usually replaced by the preproleader of S. cerevisiae alfa mating factor (MFα1). However, in most cases, only basal enzyme levels are obtained. During directed evolution in S. cerevisiae of laccases fused to the α-factor preproleader, we demonstrated that mutations accumulated in the signal peptide notably raised enzyme secretion. Here we describe different protein engineering approaches carried out to enhance the laccase activity detected in the liquid extracts of S. cerevisiae cultures. We demonstrate the improved secretion of native and engineered laccases by using the fittest mutated α-factor preproleader obtained through successive laccase evolution campaigns in our lab. Special attention is also paid to the role of protein N-glycosylation in laccase production and properties, and to the introduction of conserved amino acids through consensus design enabling the expression of certain laccases otherwise not produced by the yeast. Finally, we revise the contribution of mutations accumulated in laccase coding sequence (CDS) during previous directed evolution campaigns that facilitate enzyme production.

Enzyme production by immobilized Phanerochaete chrysosporium using airlift reactor

Biologia ◽  
2014 ◽  
Vol 69 (11) ◽  
Author(s):  
Denisse González-Ramírez ◽  
Claudia Muro-Urista ◽  
Ainhoa Arana-Cuenca ◽  
Alejandro Téllez-Jurado ◽  
Aldo González-Becerra
Keyword(s):  
Phanerochaete Chrysosporium ◽  
Peroxidase Activity ◽  
Manganese Peroxidase ◽  
Enzyme Production ◽  
Laccase Activity ◽  
Alginate Beads ◽  
Airlift Reactor ◽  
Airlift Bioreactor ◽  
Brilliant Blue ◽  
Maximum Percentage

AbstractEnzyme production by immobilized Phanerochaete chrysosporium was evaluated in airlift bioreactor and agitated cultures. Free mycelium and immobilized mycelium on alginate beads were tested in the decolourization of 50 and 500 mg/L of Remazol Brilliant Blue R. Dye concentration did not inhibit the fungi development in all tests. In addition, high decolourization percentage of dye was found with free mycelium (99%) in agitated flasks and with immobilized mycelium in airlift (98%). However, decolourization period by immobilized mycelium (120 h) was greater than that by the free mycelium (14 h). Important manganese peroxidase, lignine peroxidase and laccase activities were identified in decolourization process. Manganese peroxidase appeared to be promoted by high dye concentrations during the treatment with immobilized mycelium, but this enzyme was not detected with free mycelium in airlift. Bioreactor prompted also laccase and lignine peroxidase actions in both tests; free mycelium registered a maximum laccase action of 31.569 × 103 U/L in 70 h, whereas immobilized mycelium registered 1.680 × 103 U/L in 170 h, while lignine peroxidase secretion by free P. chrysosporium was higher (1.300 × 103 U/L) than immobilized mycelium (1.250 × 103 U/L). Maximum laccase activity coincided with the maximum percentage of decolourization, however, high peroxidase activity was identified from the start of dye treatment.

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