POLYPROTEIN PROCESSING IN ECHOVIRUS-22 - A FIRST ASSESSMENT

The major steps of the polyprotein processing of Echovirus 22 (EV22), a highly unusual member of the picornavirus family, have been characte rized for the first time by employing in vitro assay systems. Cell-fre e expression of a P1-2ABC precursor as well as VP1-2A yielded autoprot eolytically inactive proteins, suggesting that the 2A region of the EV 22 polyprotein does not contain a proteolytic activity. The intra- and intermolecular cleavage specificity of proteinase 3C, the major prote olytic enzyme in picornaviruses, was studied by expressing the enzyme of EV22 in a bacterial system as well as in the framework of precursor molecules generated by in vitro transcription/translation in a cell-f ree system. A VP1-2A precursor could very efficiently be cleaved in tt ans by the recombinant 3C, whereas the junction between P2 and P3 rem ained uncleaved. Expression of the complete P3-region in the cell-free system led to the autocatalytic release of large amounts of p22, a pr otein of the predicted molecular weight of the EV22 proteinase 3C that nas recognized by an antibody raised against the recombinant enzyme. (C) 1995 Academic Press, Inc.