![]() ![]() It showed the conservation of known consensus sequences –CreA, Hap2-3-4, PacC …–, and the existence of a particular sequence –CCTGA– which is similar to that already found to be specific of pectinolytic gene in Aspergillus, CCCTGA. In addition to the determination of transcription start site, the promoter sequence from the pnl gene was analysed. occitanis: it exhibits the highest nucleotide homology with the pnlA of Aspergillus niger but the positions of its 4 introns is completely identical to that of A. A streaking future was found in pnl gene of P. From several isolated clones, the nucleotide sequence of the pectin lyase gene was completed and led to the identification of introns and promoter–terminator regions. A genomic bank from Penicillium occitanis fungus is constructed and screened by previously isolated cDNA probe of a pectin lyase. The regulatory cis elements of fungal pectinases are well studied in Aspergillus genera but little is known in other fungal species. In conclusion HZ is a pectin lyase-overproducing hybrid with potential applications in the pectin industry. However, addition of 2 RFU and 20 U of endo- and exo-polygalacturonase, respectively, induced the hydrolysis of 92% of orange peel solids. Pectin lyase was able to hydrolyze 56% of orange peel biomass. Pectin lyase produced by the hybrid HZ was partially purified and used for the hydrolysis of orange peel. ![]() Hybrid HZ showed an increase of 450% and 1300% in pectin lyase production compared with that of A. Prototrophic segregants showed different isoenzymatic profiles and produced increased levels of pectin lyase in cultures containing lemon peel as a sole carbon source. In the present study hybrids were obtained by protoplast fusion between mutant pectinolytic Aspergillus flavipes and Aspergillus niveus CH-Y-1043 strains. Genetic improvement of pectin lyase-overproducing strains is still necessary to improve industrial processes based on this enzyme. Pectin lyases cleave the internal glycosidic bonds of pectin by β-elimination, producing non-saturated galacturonic oligomers. A purified pectin lyase may be useful in biotechnological applications such as the food industry where the liberation of toxic methanol in pectin degradation should be avoided. The optimum pH and temperature for the recombinant enzyme are 6.0 and 50 ☌, respectively, and it is stable up to 50 ☌ when exposed for 3 h. The enzyme showed no activity on polygalacturonic acid and pectin from citrus fruit 8 % esterified. It is active on highly esterified pectin, and decreases 40 % the viscosity of pectin with a degree of esterification ≥85 %. Pectin lyase has a molecular mass of 45 kDa as determined by SDS-PAGE. The recombinant enzyme was purified to homogeneity and characterized. The gene is 1334 pb long, has three introns and codes for a protein of 376 amino acid residues. In this work, the gene and cDNA of a pectin lyase from Penicillium purpurogenum have been sequenced, and the cDNA has been expressed in Pichia pastoris. This enzyme acts on highly esterified homogalacturonan, catalysing the cleavage of α-(1→4) glycosidic bonds between methoxylated residues of galacturonic acid by means of β-elimination, with the formation of 4,5-unsaturated products. Among the enzymes which degrade pectin (pectinases) is pectin lyase (E.C. The ‘hairy’ region is more complex, containing xylogalacturonan and rhamnogalacturonans I and II. The ‘smooth’ region (homogalacturonan) is a linear polymer of galacturonic acid residues with α-(1→4) linkages, substituted by methyl and acetyl residues. Pectin is composed of two basic structures: a ‘smooth’ region and a ‘hairy’ region. One of lignocellulose constituents is pectin. Lignocellulose is the major component of plant cell walls and it represents a great source of renewable organic matter. ![]()
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