Spodoptera frugiperda (Sf9) cells are widely employed for high-level expression of heterologous recombinant genes from baculovirus vectors. Using a plasmid library encoding cDNA of Sf9 cells we have identified here the Spodoptera frugiperda analog of the proprotein convertase furin which plays an important role in posttranslational protein processing. Spodoptera frugiperda furin (Sfurin) is closest related to Drosophila melanogasterfurin with which it shares an extended cysteine-rich domain, whereas mammalian furin shows high homology only in the catalytic domain. Mammalian furin and Sfurin were further compared by expression from baculovirus vectors. Substrate specificity and inhibitor profiles are identical for Sfurin and mammalian furin, whereas calcium-dependence, pH-optimum, and thermostability differ. Cleavage of recombinant influenza virus hemagglutinin was significantly enhanced in Sf9 cells after overexpression of Sfurin.