NUCLEOTIDE-SEQUENCE AND EXPRESSION IN ESCHERICHIA-COLI OF CDNAS ENCODING PAPAYA PROTEINASE OMEGA FROM CARICA-PAPAYA

We have cloned and sequenced two similar, but distinct, cDNAs from bot h fruit and leaf tissues of Carica papaya. The C-terminal portion of t he predicted amino acid (aa) sequence of one of the clones has complet e identity with the mature enzyme sequence of the cysteine proteinase papaya proteinase omega (PpOMEGA). The second clone contains ten indiv idual bp changes compared with the first and encodes a protein with th ree single-aa substitutions, only one of which is located in the matur e sequence, but most noticeably carries an additional 19-aa C-terminal extension. The clones encode pre-pro precursor isoforms of PpOMEGA. T he former of these clones has been expressed in Escherichia coli using a T7 polymerase expression system to produce insoluble pro-enzyme whi ch has been solubilized and refolded to yield auto-activable pro-PpOME GA.