Abstract
K99 pili from bovine strain B44 ofEscherichia coli were incubated in a solution containing 1% sodium dodecyl sulfate (SDS), 4M urea, 1% 2-mercaptoethanol (2-ME), and 10% glycerol at room temperature. After 2 weeks, there was a partial conversion of the pilin from its apparent molecular weight of 17, 000 to 21, 000 due to change in conformation as studied by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). After 10–12 weeks, this change in conformation was complete. The presence of 2-ME in the incubation mixture was essential for this change; this suggested the involvement of a disulfide bond. The role of the disulfide bond could also be shown by reduction and carboxamidation of the pilin. Other modifications such as performic acid oxidation, treatment with iodoacetic acid at acid pH, and 2-hydroxy-5-nitrobenzyl bromide showed that only performic acid oxidation and HNBB modification prevented change in conformation. From these data, it was concluded that the disulfide bond and tryptophan residues of the pilin are essential for the change in conformation in SDS.
Published Version
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