Auiewiriyanukul W.Saburi W.Mori H.Arthan D.Tharamak S.Mahidol University2026-03-272026-03-272026-03-17ACS Omega Vol.11 No.10 (2026) , 15795-15808https://repository.li.mahidol.ac.th/handle/123456789/115840The α-glucosidase (AaMalI) in Aedes aegypti saliva belongs to glycoside hydrolase family 13, subfamily 17 (GH13_17) and plays a crucial role in the digestion of sucrose, which is the main sugar involved in insect metabolism. The amino-acid residues in the conserved region II have been reported as the key residues for sucrose specificity in GH13_17. Using mutagenesis, this study expressed and purified recombinant AaMalI and determined the molecular mechanism related to substrate specificity. The optimal activity was at pH 6.3 and 40 °C. AaMalI had a trisaccharide specificity similar to GH13 α-glucosidases and preference for sucrose over maltose. The single mutation of Y223H and the double mutation of P222N/Y223H altered the substrate preference from sucrose to maltose. Structural analysis of the AaMalI model obtained by superimposition with the maltose-bound complex suggested that Tyr292 stabilizes the d-glucosyl moiety at subsite +1, whereas His223 indirectly contributes to maltose hydrolysis. These findings provide structural insights into AaMalI substrate specificity and support its potential as a target for vector mosquito control.Chemical EngineeringChemistryArticleSCOPUS10.1021/acsomega.5c081802-s2.0-10503300712124701343