Qualitative and quantitative assays for flagellum-mediated chemotaxis.

A primary driving force during bacterial evolution was the capacity to access compounds necessary for growth and survival. Since the species of the genus Pseudomonas are characterized by metabolic versatility, these bacteria have developed chemotactic behaviors towards a wide range of different compounds. The specificity of a chemotactic response is determined by the chemoreceptor, which is at the beginning of the signaling cascade and to which chemoattractants and chemorepellents bind. The number of chemoreceptor genes of Pseudomonas species is significantly higher than the average number in motile bacteria. Although some of the receptors have been annotated with a function, the cognate signal molecules for the majority of them still need to be identified. Different qualitative and quantitative methods are presented that can be used to study flagellum-mediated taxis.

Characterization of molecular interactions using isothermal titration calorimetry.

Isothermal titration calorimetry (ITC) is based on a simple titration of one ligand with another and the small heat changes caused by the molecular interaction are detected. From one ITC experiment the complete set of thermodynamic parameters of binding including association and dissociation constants as well as changes in enthalpy, entropy, and free energy can be derived. Using this technique almost any type of molecular interaction can be analyzed. Both ligands are in solution, and there is no need for their chemical derivatization. There are no limits as to the choice of the analysis buffer, and the analysis temperature can be set between 4 and 80 °C. This technique has been primarily applied to study the interaction between various proteins of Pseudomonas with small molecule ligands. In addition, ITC has been used to study the binding of Pseudomonas proteins to target DNA fragments.