Detection and sequence analysis of an altered pectate lyase gene in Pseudomonas syringae pv. glycinea and related bacteria.
Pectate lyase (PL) is a potent cell wall-degrading enzyme known to play a role in the microbial infection of plants. We re-examined the pectolytic property of seven representative pathovars of Pseudomonas syringae. None of the 10 P. syringae pv. glycinea strains examined exhibited pectolytic activity. However, the PL gene (pel) was detected by Southern hybridization in four out of four P. syringae pv. glycinea strains examined. A P. syringae pv. glycinea pel gene was cloned, sequenced, and predicted to encode a protein sharing 70%-90% identity in amino acid sequence with PLs produced by pectolytic pseudomonads and xanthomonads. A series of amino acid and nucleotide sequence analyses reveal that (i) the predicted P. syringae pv. glycinea PL contains two regions in the amino acid sequence that may affect the formation of a beta-helix structure important for the enzyme activity, and (ii) the P. syringae pv. glycinea pel gene contains a single-base insertion, a double-base insertion, and an 18-bp deletion, which can lead to the synthesis of an inactive PL protein. The function of P. syringae pv. glycinea PL could be restored by removing the unwanted base insertions and by filling in the 18-bp deletions by site-directed mutagenesis. The altered pel sequence was also detected by polymerase chain reaction and nucleotide sequencing in the genomes of other pathovars of P. syringae, including phaseolicola and tagetis.
Liao
Ching-Hsing
CH
Eastern Regional Research Center, Agricultural Research Center, United States Department of Agriculture, Wyndmoor, PA 19038, USA. cliao@errc.ars.usda.gov
Fett
William
W
Tzean
Sean-Shong
SS
Hoffman
Gabriel
G
D008969:Molecular Sequence Data; D011133:Polysaccharide-Lyases / Q000737:chemistry / Q000235:genetics*; D044224:Pseudomonas syringae / Q000235:genetics*; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D012689:Sequence Homology, Nucleic Acid
D011133:Polysaccharide-Lyases; C023911:pectate lyase