Erwinia carotovora subsp. carotovora (Ecc) strain EC14, a Gram-negative
bacterium, causes soft rot on several crops, including potato.
Maceration of potato tuber tissue is caused by secreted pectolytic
enzymes. Other cell-degrading enzymes may also have roles in
pathogenesis, including cellulases, phospholipases, and protease(s).
The objectives of this research were to (1) characterize Ecc extracellular protease (Prt) and (2) elucidate its role in potato soft
rot. A gene encoding a Prt, prt1, was cloned from cosmid pCA7
containing Ecc genomic DNA into plasmid pSK1. Escherichia coli
transformed with pSK1 or pSK23, a subclone of pSK1, produced
intracellularly a 38 kDa Prt with the same pI (4.8) as the secreted Ecc
Prt. Prt1 activity produced by E.coli/pSK23 was inhibited by phenanthroline, which inhibits Zn-metalloproteases, but not by Ecc
intracellular proteins. Analysis of deletion mutants indicated a 1.2 kb
region necessary for Prt1 production. Sequencing of the pSK1 insert
revealed a 1,041 bp open reading frame (ORF1) corresponding to the prtl
region. ORF1 encodes a putative polypeptide of 347 amino acids with a
total molecular mass of 38.8 kDa.
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