Foot-and-mouth disease virus leader proteinase - Involvement of C-terminalresidues in self-processing and cleavage of eIF4GI

The leader proteinase (L-pro) of foot-and-mouth disease virus frees itself from the nascent polyprotein, cleaving between its own C terminus and the N terminus of VP4 at the sequence Lys-Leu-Lys- down arrow -Gly-Ala-Gly. Subs equently, the L-pro impairs protein synthesis from capped mRNAs in the infe cted cell by processing a host protein, eukaryotic initiation factor 4GI, a t the sequence Asn-Leu-Gly- down arrow -Arg-Thr-Thr. A rabbit reticulocyte lysate system was used to examine the substrate specificity of L-pro and th e relationship of the two cleavage reactions. We show that L-pro requires a basic residue at one side of the scissile bond to carry out efficient self -processing. This reaction is abrogated when leucine and lysine prior to th e cleavage site are substituted by serine and glutamine, respectively. Howe ver, the cleavage of eIF4GI is unaffected by the inhibition of self-process ing. Removal of the 18-amino acid C-terminal extension of Lpr, slowed eIF4G I cleavage; replacement of the C-terminal extension by unrelated amino acid sequences further delayed this cleavage. Surprisingly, wild-type L-pro and the C-terminal variants all processed the polyprotein cleavage site in an intermolecular reaction at the same rate. However, when the polyprotein cle avage site was part of the same polypeptide chain as the wild-type Lb(pro), the rate of processing was much more rapid. These experiments strongly sug gest that self-processing is an intramolecular reaction.