Transglutaminase　(TGase)　induced-crosslinking　of　soy　protein　isolate　(SPI)　was　markedly　influenced　by　the　substrate　aggregation　state.　Results　showed　that　appropriate　heating　significantly　accelerated　the　TGase　crosslinking,　and　the　7S　and　11S　acidic　subunits　were　more　susceptible　to　the　enzyme　than　the　11S　basic　proteins.　The　content　of　ε-(γ-glutamyl)-lysine　isopeptide　bonds　increased　from　4.74　to　8.61　μmol/g　protein　when　the　heating　intensity　was　increased　from　75　°C　for　15　min　to　95　°C　for　30　min,　due　to　sufficient　unfolding　of　the　protein　structure.　Rheological　data　indicated　that　the　gel　formed　from　the　SPI　heated　at　95　°C　for　30　min　exhibited　the　best　properties,　with　a　60　%　increase　in　the　storage　modulus　compared　with　the　unheated　sample.　However,　excessive　heating　(95　°C,　60–120　min)　caused　severe　aggregation　of　SPI　and　formation　of　insoluble　aggregates,　resulting　in　poor　crosslinking　efficiency　and　weaker　gel　properties.　©　2024　Elsevier　Ltd