Modification of histidine residues by diethyl pyrocarbonate leads to inactivation of the Rhodospirillum rubrum RrF₁-ATPase

The RrF₁-ATPase from the photosynthetic bacterium Rhodospirillum rubrum was rapidly and completely inactivated by diethyl pyrocarbonate (DEPC) at pH 6.0 and 22°C. When applied in 1 step 5 mM DEPC were required for > 90% inactivation and this DEPC-modified enzyme showed an increase in absorption at 242 nm as well as a decrease in absorption at 280 nm, suggesting modification of both histidine and tyrosine residues. Complete inactivation of the RrF₁-ATPase could be obtained with only 250 μM DEPC when applied in 5 separate steps of 50 μM each. The only absorption change observed under these conditions was an increase at 242 nm indicating that the inactivation can be correlated with modification of histidine residues. Complete inactivation requires the modification of 2-3 histidine residues per molecule of RrF₁.