Chemoreceptors in Escherichia coli are coupled to the flagella by a labile phosphorylated intermediate, CheY approximately P. Its activity can be inferred from the rotational bias of flagellar motors, but motor response is stochastic and limited to a narrow physiological range. Here we use fluorescence resonance energy transfer to monitor interactions of CheY approximately P with its phosphatase, CheZ, that reveal changes in the activity of the receptor kinase, CheA, resulting from the addition of attractants or repellents. Analyses of cheR and/or cheB mutants, defective in receptor methylation/demethylation, show that response sensitivity depends on the activity of CheB and the level of receptor modification. In cheRcheB mutants, the concentration of attractant that generates a half-maximal response is equal to the dissociation constant of the receptor. In wild-type cells, it is 35 times smaller. This amplification, together with the ultrasensitivity of the flagellar motor, explains previous observations of high chemotactic gain.