PMID:15037246

Citation	Okuno, T, Yamada-Inagawa, T, Karata, K, Yamanaka, K and Ogura, T Spectrometric analysis of degradation of a physiological substrate sigma32 by Escherichia coli AAA protease FtsH. J. Struct. Biol. 146:148-54
Abstract	We have established a fluorescence polarization assay system by which degradation of sigma32, a physiological substrate, by FtsH can be monitored spectrometrically. Using the system, it was found that an FtsH hexamer degrades approximately 0.5 molecules of Cy3-sigma32 per min at 42 degrees C and hydrolyzes approximately 140 ATP molecules during the degradation of a single molecule of Cy3-sigma32. Evidence also suggests that degradation of sigma32 proceeds from the N-terminus to the C-terminus. Although FtsH does not have a robust enough unfoldase activity to unfold a tightly folded proteins such as green fluorescent protein, it can unfold proteins with lower T(m)s such as glutathione S-transferase (T(m) = 52 degrees C).
Links	PubMed Online version:10.1016/j.jsb.2003.10.019
Keywords	ATP-Dependent Proteases; Bacterial Proteins/isolation & purification; Bacterial Proteins/metabolism; Carbocyanines; Escherichia coli Proteins/metabolism; Fluorescence Polarization; Glutathione Transferase; Heat-Shock Proteins/genetics; Heat-Shock Proteins/metabolism; Membrane Proteins/isolation & purification; Membrane Proteins/metabolism; Mutation, Missense; Recombinant Fusion Proteins; Sigma Factor/genetics; Sigma Factor/metabolism; Spectrometry, Fluorescence/methods
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