Characterization of Erwinia chrysanthemi PY35 cel and pel gene existing in tandem and rapid identification of their gene products

Abstract

Genomic DNA of the phytopathogenic Erwinia chrysanthemi PY35 was partially digested with Sau3AI, ligated into the BamHI site of pBluescript II SK+, and introduced into E. coli. One clone that was able to hydrolyse carboxymethylcellulose and polygalacturonic acid was selected. A 2.9 kb fragment containing the pelL1 gene (pPY300) and cel5Z gene (pPY401) in tandem was subcloned and sequenced. The pelL1 and cel5Z genes had open reading frames of 1278 bp and 1281 bp encoding 425 and 426 amino acid residues with calculated molecular weights of 45,649 Da and 46,473 Da, respectively. pelL1 and cel5Z carried a typical prokaryotic signal peptide of 24 and 41 amino acid residues, respectively. The apparent molecular masses of the proteins when expressed in E. coli cells were approximately 43 kDa (PelL1) and 42 kDa (Cel5Z) as assessed by PGA-SDS-PAGE and CMC-SDS-PAGE. (C) 2000 Academic Press.