Universal Stress Protein Regulates Electron Transfer and Superoxide Generation Activities of the Cytochrome bc1 Complex from Rhodobacter sphaeroides.

Abstract

Interactions between Rhodobacter sphaeroides cytochrome bc1 complex (Rsbc1) and soluble cytosolic proteins were studied by a precipitation pull-down technique. After being purified, detergent-dispersed Rsbc1 complex was incubated with soluble cytosolic fraction and then dialyzed in the absence of detergent; the interacting proteins were coprecipitated with Rsbc1 complex upon centrifugation. One of the cytosolic proteins pulled down by Rsbc1 complex was identified by liquid chromatography-coupled tandem mass spectrometry (LC/MS/MS) to be the reported R. sphaeroides universal stress protein (UspA). Incubating purified UspA with the detergent dispersed bc1 complex resulted in an increase in the Rsbc1 complex activity by 60% and a decrease in superoxide generation activity by the complex by more than 70%. These UspA effects were only observed with Rsbc1 complexes containing subunit IV and assayed under aerobic conditions. These results suggest that the interaction between UspA and Rsbc1 complex may play an important role in R. sphaeroides cells during oxidative stress. Using a biotin label transfer technique, cytochrome c1 of the Rsbc1 complex was identified as the interacting site for UspA.