Haykır, Nazife IşıkKosaoğlu, HacimTuncel, AliÇelebi, Serdar Suut2026-04-012026-04-012026Haykır, N. I., Kosaoğlu, H., Tuncel, A., & Çelebi, S. S. (2026). Thermo, pH, and ionic-strength-responsive MPEG–PEI copolymer: A reversible soluble–insoluble support promotes glucoamylase immobilization and sustainable catalysis. Biocatalysis and Agricultural Biotechnology, 73, pp. 1-12. https://doi.org/10.1016/j.bcab.2026.1039871878-8181https://doi.org/10.1016/j.bcab.2026.103987https://hdl.handle.net/20.500.13055/1373Glucoamylase (GA) plays a crucial role in the saccharification of starch and other related oligo saccharides in the food and fermentation industries. The construction of an immobilized GA using a reversibly soluble/insoluble methoxy polyethylene glycol–polyethyleneimine (MPEG–PEI) copolymer synthesized via carbodiimide chemistry is presented in this work. Various MPEG:GA ratios were assessed to optimize enzyme loading and catalytic activity. Immobilized GA retained up to 12.6% of its theoretical activity but demonstrated excellent operational stability and reusability. Substrate-assisted immobilization, using maltose and maltodextrin to protect GA's active site during synthesis of the carrier, significantly improved enzyme activity—up to a 3.1- fold increase compared to the unprotected system. While maltodextrin yielded higher initial activity, maltose with a lower initial activity offered better long-term stability, maintaining 70% of its initial activity even after 11 reuse cycles. Kinetic analysis using the Lambert-W function revealed increased Km for the immobilized enzyme, indicating decreased substrate affinity due to a possible conformational change on the immobilized form of GA. While a minor decrease in conversion efficiency was observed under high initial substrate concentrations in practice, the reversibly soluble/insoluble MPEG–PEI copolymer exhibited effective thermo-responsive char acteristics, enabling the sustained and recyclable use of GA in starch hydrolysis and offering considerable potential for other future biotechnological applications.eninfo:eu-repo/semantics/closedAccessGlucoamylaseImmobilizationThermo-Responsive PolymerBiocatalysisEnzyme StabilityArticle10.1016/j.bcab.2026.10398773112Q22-s2.0-105031378847Q1