The trp repressor of Escherichia coli (TR), although generally considered to be dimeric, has been shown by fluorescence anisotropy of extrinsically labeled protein to undergo oligomerization in solution at protein concentrations in the micromolar range (Fernando, T., and C. A. Royer 1992. Biochemistry. 31:3429–3441). Providing evidence that oligomerization is an intrinsic property of TR, the present studies using chemical cross-linking, analytical ultracentrifugation, and molecular sieve chromatography demonstrate that unmodified TR dimers form higher order aggregates. Tetramers and higher order species were observed in chemical cross-linking experiments at concentrations between 1 and 40 microM. Results from analytical ultracentrifugation and gel filtration chromatography were consistent with average molecular weight values between tetramer and dimer, although no plateaus in the association were evident over the concentration ranges studied, indicating that higher order species are populated. Analytical ultracentrifugation data in presence of corepressor imply that corepressor binding destabilizes the higher order aggregates, an observation that is consistent with the earlier fluorescence work. Through the investigation of the salt and pH dependence of oligomerization, the present studies have revealed an electrostatic component to the interactions between TR dimers.
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