Analysis of conductance responses during depolymerization of pectate by soft rot Erwinia spp. and other pectolytic bacteria isolated from potato tubers.
Different bacteria isolated from potato tubers were screened for their pectolytic properties by examining pitting in polypectate agar, recording conductance responses in polypectate medium and performing potato tuber soft rot tests. For bacteria found positive in conductimetry, the role of polygalacturonase (PG) and pectate lyase (PL) in the generation of conductance changes in a polygalacturonic acid (PGA) medium was further analysed using enzyme activity staining after gel electrophoresis and high-performance anion exchange chromatography. The extent of the conductance changes during depolymerization of PGA was dependent on the amounts of galacturonate monomers and oligomers accumulated in the medium. In comparison with an unidentified saprophyte and a Klebsiella strain, both mainly having PL activity, soft rot Erwinia spp. rapidly produced larger conductance responses, due to a combined action of multiple forms of PG and PL. The responses of Erwinia spp. were initially associated with the accumulation of large amounts of monomers and saturated dimers to heptamers, due to PG activity. Subsequently, as well as monomers and saturated dimers, large amounts of unsaturated dimers were also detected, due to PL activity. The role of PG as an important conductimetric factor was also demonstrated for a pectinase preparation derived from Aspergillus niger. Besides detection, automated conductimetric assays in pectate media may also be useful for monitoring of pectolytic activity in pectinase preparations and for screening of pectolytic activity of microorganisms under different media and growth conditions.
Fraaije
B A
BA
DLO-Centre for Plant Breeding and Reproduction Research (CPRO-DLO), Wageningen, The Netherlands.
Bosveld
M
M
Van den Bulk
R W
RW
Rombouts
F M
FM
D002852:Chromatography, Ion Exchange; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D004885:Erwinia / Q000201:enzymology*; D010368:Pectins / Q000378:metabolism*; D010935:Plant Diseases / Q000382:microbiology*; D011096:Polygalacturonase / Q000378:metabolism; D011108:Polymers / Q000378:metabolism; D011133:Polysaccharide-Lyases / Q000378:metabolism; D012967:Sodium Dodecyl Sulfate; D011198:Solanum tuberosum / Q000382:microbiology*
D011108:Polymers; D012967:Sodium Dodecyl Sulfate; D010368:Pectins; D011096:Polygalacturonase; D011133:Polysaccharide-Lyases; C023911:pectate lyase; C003181:polygalacturonic acid