FOOT-AND-MOUTH-DISEASE VIRUS LEADER PROTEINASE - PURIFICATION OF THE LB FORM AND DETERMINATION OF ITS CLEAVAGE SITE ON EIF4-GAMMA

Many picornaviruses cause a dramatic decrease in the translation of ce llular mRNAs in the infected cell, without affecting the translation o f their own RNA. Specific proteolysis of protein synthesis initiation factor eIF-4 gamma occurs during infection with rhinoviruses, enterovi ruses, and aphthoviruses, apparently leading to an inability of the ri bosomes to bind capped mRNAs. Cleavage of eIF-4 gamma in human rhinovi ruses and enteroviruses is carried out by the viral 2A proteinase; in aphthoviruses (i.e., foot-and-mouth disease viruses), the leader prote inase is responsible for this reaction. We describe here the purificat ion to homogeneity of the Lb form of the leader proteinase expressed i n Escherichia coli. The primary cleavage products of eIF-4 gamma obtai ned in vitro with purified leader or 2A proteinase are electrophoretic ally indistinguishable from those found during infection in vivo. Howe ver, additional proteolysis products of elF-4 gamma are observed with the leader proteinase and the human rhinovirus type 2 2A proteinase in vitro. The cleavage site of the leader proteinase in eIF-4 gamma from rabbit reticulocyte was determined by sequencing the purified C-termi nal cleavage product by automated Edman degradation. The cleavage site is between Gly-479 and Arg-480 and thus differs from that of rhinovir us and enterovirus 2A proteinases, which cleave between Arg-486 and Gl y-487.