Purification, characterization and three-dimensional structure prediction of multicopper oxidase Laccases from Trichoderma lixii FLU1 and Talaromyces pinophilus FLU12

Abstract Broad-spectrum biocatalysts enzymes, Laccases, have been implicated in the complete degradation of harmful pollutants into less-toxic compounds. In this study, two extracellularly produced Laccases were purified to homogeneity from two different Ascomycetes spp. Trichoderma lixii FLU1 (TlFLU1) and Talaromyces pinophilus FLU12 (TpFLU12). The purified enzymes are monomeric units, with a molecular mass of 44 kDa and 68.7 kDa for TlFLU1 and TpFLU12, respectively, on SDS-PAGE and zymogram. It reveals distinct properties beyond classic protein absorption at 270–280 nm, with TlFLU1's peak at 270 nm aligning with this typical range of type II Cu site (white Laccase), while TpFLU12's unique 600 nm peak signifies a type I Cu 2+ site (blue Laccase), highlighting the diverse spectral fingerprints within the Laccase family. The K m and k cat values revealed that ABTS is the most suitable substrate as compared to 2,6-dimethoxyphenol, caffeic acid and guaiacol for both Laccases. The bioinformatics analysis revealed critical His, Ile, and Arg residues for copper binding at active sites, deviating from the traditional two His and a Cys motif in some Laccases. The predicted biological functions of the Laccases include oxidation–reduction, lignin metabolism, cellular metal ion homeostasis, phenylpropanoid catabolism, aromatic compound metabolism, cellulose metabolism, and biological adhesion. Additionally, investigation of degradation of polycyclic aromatic hydrocarbons (PAHs) by purified Laccases show significant reductions in residual concentrations of fluoranthene and anthracene after a 96-h incubation period. TlFLU1 Laccase achieved 39.0% and 44.9% transformation of fluoranthene and anthracene, respectively, while TpFLU12 Laccase achieved 47.2% and 50.0% transformation, respectively. The enzyme structure–function relationship study provided insights into the catalytic mechanism of these Laccases for possible biotechnological and industrial applications

Soil microbes and associated extracellular enzymes largely impact nutrient bioavailability in acidic and nutrient poor grassland ecosystem soils

Abstract Understanding the role of soil microbes and their associated extracellular enzymes in long-term grassland experiments presents an opportunity for testing relevant ecological questions on grassland nutrient dynamics and functioning. Veld fertilizer trials initiated in 1951 in South Africa were used to assess soil functional microbial diversity and their metabolic activities in the nutrient-poor grassland soils. Phosphorus and liming trials used for this specific study comprised of superphosphate (336 kg ha−1) and dolomitic lime (2250 kg ha−1) (P + L), superphosphate (336 kg ha−1) (+ P) and control trials. These soils were analyzed for their nutrient concentrations, pH, total cations and exchange acidity, microflora and extracellular enzyme activities. The analysed soil characteristics showed significant differences except nitrogen (N) and organic carbon (C) concentrations showing no significant differences. P-solubilizing, N-cycling and N-fixing microbial diversity varied among the different soil treatments. β-glucosaminidase enzyme activity was high in control soils compared to P-fertilized and limed soils. Alkaline phosphatase showed increased activity in P-fertilized soils, whereas acid phosphatase showed increased activity in control soils. Therefore, the application of superphosphate and liming influences the relative abundance of bacterial communities with nutrient cycling and fixing functions which account for nutrient bioavailability in acidic and nutrient stressed grassland ecosystem soils