Oxygen and redox sensing by two-component systems that regulate behavioral responses: behavioral assays and structural studies of aer using in vivo disulfide cross-linking.

摘要: A remarkable increase in the number of annotated aerotaxis (oxygen-seeking) and redox taxis sensors can be attributed to recent advances bacterial genomics. However, silico predictions should supported by behavioral assays genetic analyses that confirm an or function. This chapter presents a collection procedures have been highly successful characterizing Escherichia coli. The methods are described enough detail enable investigators other species adapt for their use. gas flow cell is used quantitate temporal responses bacteria step decrease oxygen partial pressure potential. Bacterial behavior spatial gradients analyzed using optically flat capillaries soft agar plates (succinate tryptone agar). We describe two approaches estimate preferred attracts species; this concentration important understanding microbial ecology. At molecular level, we determine structure topology Aer, membrane receptor aerotaxis. Cysteine-scanning mutagenesis vivo disulfide cross-linking utilize oxidant Cu(II)-(1,10-phenanthroline)(3) bifunctional sulfhydryl-reactive probes. Finally, boundaries transmembrane segments receptors such as Aer. These include 5-iodoacetamidofluorescein, 4-acetamido-4-disulfonic acid, disodium salt (AMS), methoxy polyethylene glycol maleimide, 5-kDa mass probe alters mobility Aer on SDS-PAGE.