Studies of E. coli cells

In an attempt to assess the biological relevance of the cluster conversion observed in the purified FNR transcription factor and the oxygen sensing properties of this protein in vivo, we have studied by Mossbauer spectroscopy several preparations of . coli maintained in anaerobic media and then submitted to controlled aerobic conditions. Our strategy was to study pairs of whole cell samples in which FNR is either over-expressed or is lacking which would enable us to recognize the contri- 

We have searched for the growth conditions for which the unresolved back- 

Our preliminary experiments on whole cells show that the molecular basis of the FNR inactivation in aerobic medimn, namely the [4Fe-4S] + cluster degradation via a [2Fe-2S] intermediate, previously deduced from studies of purified FNR, is valid also in vivo. It is not yet clear whether the 4Fe 2Fe cluster conversion observed in vitro as well as in vivo puts the protein in a ready state from which the 4Fe cluster can be rebuilt once the bacterial environment becomes anaerobic again. Conceivably, the [2Fe-2S] cluster may be just an intermediate in the pathway of complete aerobic degradation of the 4Fe-4S cluster. These are questions which need to be addressed by future studies. 

The present study demonstrates that the Mossbauer spectral signature of the FNR protein is readily recognizable in spectra of whole E. coli cells, provided this protein is over-expressed. Moreover, the approach used here may be applicable for probing other proteins in their native cellular environment. 

This work was supported by the United States Public Health Service, National Institutes for Heal A grants GM-45844 (to PJK) and GM-22701 (to EM). PJK was also a recipient of a Young Investigator Award from the National Science Foundation and the Shaw Scientist Award from the Milwaukee Foundation. 

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