INTEGRATED IN SILICO STRUCTURAL AND FUNCTIONAL CHARACTERIZATION OF CELLULASE ENZYMES FROM CELLULOLYTIC YEASTS

Abstract

Yeasts are microscopically eukaryotic fungi that are distributed everywhere in nature, but are most abundant in environments with rich sugar content, such as fruits and flower nectar; over 1,500 species of yeasts have been found, and one of them, Saccharomyces cerevisiae, is a model organism that has been most intensely studied. Few yeasts are now found to produce cellulases. An integrated in silico approach was employed to comparatively characterize cellulase enzymes from the isolates Vanrija humicola, Geotrichum candidum, and Aureobasidium pullulans, focusing on their structural, functional, and regulatory attributes. Protein sequences were retrieved from UniProt and assessed for homology by BLAST. Amino acid composition, molecular weight, and basic physicochemical properties, instability index, molar extinction coefficient, and the prediction of secondary structural elements were performed. Functional domains, conserved motifs, phosphorylation sites, transmembrane regions, and salt-bridge compositions were identified to evaluate protein functionality and stability. Physicochemical parameters were assessed by ProtParam, while signal peptides and transmembrane helices were predicted using the DeepTMHMM programs, respectively. Three-dimensional modeling of structures was performed using SWISS-MODEL and trRosetta, and model quality was further validated by QMEAN, and InterPro analyses. Other analyses included motif scanning, phosphorylation site prediction by means of NetPhos. The in-silico characterization described here is useful in understanding structure-function relationships in yeast proteins, it therefore supports further applications in biotechnology and industrial research.Sequence homology and evolutionary analyses revealed strong conservation of catalytic residues and motifs typical of endo-β-1,4-glucanases, indicating a shared mechanism of cellulose hydrolysis. Overall, these findings underscore the functional complementarity of fungal cellulases, integrating structural robustness with catalytic flexibility. The distinct yet synergistic properties of the analyzed enzymes support their relevance as promising candidates for targeted applications in lignocellulosic biomass conversion, bioethanol production, and industrial biotechnology, while providing a rational framework for future experimental validation and enzyme engineering strategies.