Enzymatic Properties and Structural Insights Into the Derhamnosylating Alkaline α‐l‐Rhamnosidase From Aspergillus flavus

Abstract α‐ l ‐Rhamnosidases are ubiquitous enzymes responsible for derhamnosylation of α‐ l ‐rhamnose moiety from a variety of glycoconjugates and numerous natural glycosides. An α‐ l ‐rhamnosidase‐secreting fungal strain was isolated from soil sample. Further, it was identified as Aspergillus flavus through internal transcribed spacer (ITS) gene sequencing. The enzyme was purified to homogeneity using ion‐exchange and gel filtration chromatography and exhibited molecular weight of 71 ± 1 kDa. The maximum catalytic efficiency for the α‐ l ‐rhamnosidase was established to be pH 10.0 and at a temperature of 50°C. The purified enzyme exhibits a K m 0.41 ± 0.06 mM and a V max 2.43 ± 0.17 µmol/min/mg for naringin hydrolysis. In this study, we modeled the 3D structure of A. flavus α‐ l ‐rhamnosidase using SWISS Model and validated it via PDBsum and PROCHECK. Molecular docking of A. flavus α‐ l ‐rhamnosidase with naringin and p ‐nitrophenyl‐α‐ l ‐rhamnopyranoside ( p NPR) identified key binding interactions. Electrostatic surface analysis highlighted ligand‐binding sites, revealing crucial residues for substrate recognition and enzyme stability. Active site residues of A. flavus α‐ l ‐rhamnosidase forming hydrogen bonds and hydrophobic interactions with naringin and p NPR were identified, providing insights into substrate specificity and its potential applications in glycoside hydrolysis.