Properties of Mutants in Galactose Taxis and Transport

Abstract

8-Methylgalactoside (mgl) permease mutants of Escherichia coli, which are defective in three genes, mglA, mglB, and mglC, were assayed for galactose taxis and galactose transport. The mglB product is the galactose-binding protein. Previous evidence, supported by our new findings, shows that the galactose-binding protein is the recognition component for galactose taxis as well as for galactose transport. Most mutants defective in mglB showed strong effects on both chemotaxis and transport; however, a couple showed effects chiefly on one process or the other, thus allowing a separation of chemotaxis and transport. The mglA and mglC products have not yet been identified, but they must be components of the galactose transport machinery since mutants defective in mglA or mglC, or both, showed strongly reduced transport. Although some of these mutants showed little chemotaxis, most gave close to wild-type chemotactic responses. Thus, transport is not required for galactose taxis. The bacteria detect changes in the fraction of binding protein associated with galactose, not changes in the rate of transport.MATERIALS AND METHODS Bacterial strains. The bacterial strains employed are listed in Tables 1 and 2 and have been further described in (17). Media. Tryptone broth is 1% tryptone, 0.5% NaCl.Vogel-Bonner minimal medium (22) was used as mineral salts base. For growth of bacteria it was supplemented with any essential amino acids (0.5 mM), thiamine (1 gg/ml), and 0.4% D-galactose or glycerol. Eosine methylene blue (EMB) galactose plates contained 2.75% EMB agar base (Difco) and 1.0% galactose.Radioactivity. D-[1-14C ]Galactose, at specific activity of 52.9 mCi/mmol was obtained from New England Nuclear Corp. Under our conditions of counting (Packard Tri-Carb liquid scintillation counter), 1 pmol was found to correspond to 50 counts/min. Preparation of galKstrains. A strain to be made galK-(defective in galactokinase) was lysogenized with X cI857 xis-1 Jam6 (gift from J. S. Parkinson). These lysogens are not viable at high temperature since the repressor is thermolabile and the phage cannot be excised (see 11; J. S. Parkinson, personal communication). Bacteria were transduced by P1 517 on August 1, 2020 by guest

Differences in signalling by directly and indirectly binding ligands in bacterial chemotaxis

“…Although such a high flow rate generally increased measurement errors, it resulted in a significant increase in response amplitudes to the lowest concentrations of some ligands when compared with a five-fold lower flow rate (Supplementary Figure S2), and led to modestly higher estimates of the effective threshold sensitivity ( Supplementary Table SI). Such reduction of the response threshold by ligand uptake is consistent with previous observations made in capillary assays (Ordal and Adler, 1974;Hazelbauer, 1975;Zhang et al, 1999), and its extent was ligand-dependent as expected from differences in the rates of ligand uptake. Only minor effects were observed for aspartate or galactose, whereas depletion effects for serine or ribose were substantially larger.…”

Chemotaxis in Bacteria

“…Thus, certain components of the transport system, and the process of transport itself, are not required for chemotaxis (at least for certain chemoreceptors). This has been studied extensively in the case of galactose, where transport is clearly not required (10,14,44). Two genes of Type 2 were found for the /3-methylgalactoside transport system for galactose (44).…”

“…This has been studied extensively in the case of galactose, where transport is clearly not required (10,14,44). Two genes of Type 2 were found for the /3-methylgalactoside transport system for galactose (44). Some of the mutations in these genes abolished transport without affecting chemotaxis; other mutations in these genes affected chemotaxis as well (44).…”

“…Two genes of Type 2 were found for the /3-methylgalactoside transport system for galactose (44). Some of the mutations in these genes abolished transport without affecting chemotaxis; other mutations in these genes affected chemotaxis as well (44). Such chemotactic defects may reflect interactions, direct or indirect, that these components normally have with the chemoreception machinery, or some kind of unusual interaction of the mutated component with the binding protein which would hinder its normal function in chemoreception.…”

Chemotaxis: Old and New

“…We now know that there are four kinds of these methylatable transducers in E. coli and mutants in each of these are recognized ( Fig. 11) (Ordal and Adler, 1974;Springer et aI., 1977;Silverman and Simon, 1977;Kondoh et aI., 1979;Hazelbauer and Harayama, 1979;Boyd et al, 1981;Manson et aI., 1986). Thanks to the laboratories of Simon Krikos et aI., 1983), Koshland (Russo and Koshland, 1983), and Hazelbauer (Bollinger et al, 1984), the complete amino acid sequence of these four proteins is now known (see my 1987 review for a comparison of the four MCP sequences determined by the above authors; Adler, 1987).…”