Question

Topic: Bacteria Chemotaxis (E. coli)

1. You are studying the interaction of CheY and the P1-P2 domains of CheA. You have used site specific mutagenesis to replace F214 of CheA with an alanine residue.

A. What genetic experimental assay could you use to determine the effect of the F214 to A214 substitution on the interaction between CheA-P1P2 and CheY. Explain how the assay works.

B. What would be the effect of the F214 to A214 substitution on the interaction between CheA and CheY?

C. You have created an E. coli strain in which the wild-type cheA gene has been deleted and he deletion strain has been transformed with a plasmid containing the cheA-A214 mutant gene. In medium that has very low concentration of serine how would the swimming behavior in the mutant strain differ from that of the wild-type strain? Explain your answer.

Notes:

Notes:
Chemotaxis in E. coli and other enteric bacteria Low signal concentration (Ser) Tsr-CheW-CheA-P — CW rotation (tumble) — CheY-P — Flagella motor Chez CheY Increasing signal concentration (Ser) Ser-Tsr-Chew-Chea Chey Flagella motor — CCW rotation (run) MW

Chemotaxis –migration towards attractants and away from repellents. (A) Bacteria such as E.

coli exhibit two modes of swimming: runs and tumbles. (B) Cells tend to continue on course

when running towards attractants; when swimming away from attractants they tend to tumble

and change direction. When flagella rotate in the counter clockwise direction (CCW) the flagella

forms a bundle and the cell moves in smooth direction (run). The tendency to rotate in a

clockwise is suppress so you get an extended run in the direction of a gradient. When the flagella

rotate in the clockwise (CW) direction, they rotate separately and the cell tumbles. The question

we will address is what are the molecular mechanism of that couple the sensing of a food source

like serine to the control of the rotation of the flagella motor.