CHARACTERIZATION OF A NEW 4-CHAIN COILED-COIL - INFLUENCE OF CHAIN-LENGTH ON STABILITY

Limited information is available on inherent stabilities of four-chain coiled-coils. We have developed a model system to study this folding motif using synthetic peptides derived from sequences contained in the tetramerization domain of Lac repressor. These peptides are tetrameri c as judged by both gel filtration and sedimentation equilibrium and t he tetramers are fully helical as determined by CD. The four-chain coi led-coils are well folded as judged by the cooperativity of thermal un folding and by the extent of dispersion in aliphatic chemical shifts s een in NMR spectra. In addition, we measured the chain length dependen ce of this four-chain coiled-coil. To this end, we developed a general procedure for nonlinear curve fitting of denaturation data in oligome ric systems. The dissociation constants for bundles that contain alpha -helical chains 21, 28, and 35 amino acids in length are 3.1 x 10(-12) 6.7 x 10(-23), and 1.0 x 10(-38) M(3), respectively. This corresponds to tetramer stabilities (in terms of the peptide monomer concentratio n) of 180 mu M, 51 nM, and 280 fM, respectively. Finally, we discuss t he rules governing coiled-coil formation in light of the work presente d here.