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Although  c -type cytochromes ( c -Cyts) mediating metal reduction have been mainly investigated with  in vitro  purified proteins of dissimilatory metal reducing bacteria, the  in vivo  behavior of  c -Cyts is still unclear given the difficulty in measuring the proteins of intact cells. Here,  c -Cyts in living  Shewanella putrefaciens  200 (SP200) was successfully quantified using diffuse-transmission UV/Vis spectroscopy due to the strong absorbance of hemes, and the  in situ  spectral kinetics of Cr(VI) reduction by  c -Cyts were examined over time. The reduced product Cr(III) observed on the cell surface may play a role in inhibiting the Cr(VI) reduction and reducing the cell numbers with high concentrations (>200 μM) of Cr(VI) evidenced by the 16S rRNA analysis. A brief kinetic model was established with two predominant reactions, redox transformation of  c -Cyts and Cr(VI) reduction by reduced  c -Cyts, but the fitting curves were not well-matched with  c -Cyts data. The Cr(III)-induced inhibitory effect to the cellular function of redox transformation of  c -Cyts was then added to the model, resulting in substantially improved the model fitting. This study provides a case of directly examining the reaction properties of outer-membrane enzyme during microbial metal reduction processes under physiological conditions.

In Situ Spectral Kinetics of Cr(VI) Reduction by c-Type Cytochromes in A Suspension of Living Shewanella putrefaciens 200