Isolation of chemotactic signalling mutants in escherichia coli

Abstract

Bacterial chemotaxis is the response of an organism to a chemical gradient. Chemotactic sensory transducers located in the cytoplasmic membrane sense the stimulus and change the flagellar rotation frequency through the cytoplasmic sensory transduction pathway. Trg is the transducer for ribose and galactorse. For the purpose of understanding the signalling domain of Trg, the trg gene on a multicopy plasmid was mutagenized by hydroxylamine in vitro. After screening on chemotactic swarm plate, three mutant plasmids which show generally nonchemotactic phenotypes were isolated. They did``nt show any chemotactic activity on multicopy plasmid but gave slight response to ribose and galactose in a single chromosomal copy like previously characterized signalling mutation trg-21. The proteins from the mutations, named trg-202 and trg-203, show abnormal mobility on SDS-PAGE resulting from the defect in the deamidation like trg-21 In contrast, another mutation trg-201 is able to undergo deamidation in the same manner as wild type Trg protein. The mutations, trg-202 and trg-203, exhibit a pronounced counterclockwise bias in flagellar rotation while trg-201 shows exclusively clockwise rotation.