Contact with a chloroform/water interface at 2 degrees C induces contraction of fd filamentous phage into rodlike I-forms; this contraction is accompanied by a decrease in the magnitude of circular dichroism spectral intensity near 222 nm and an increase near 210 nm. Comparisons with circular dichroism spectra of 100% helical poly-L-lysine and N-bromosuccinimide-oxidized fd phage indicate that the spectral change accompanying the fd to I-forms transition is due primarily to a change in the contributions from the single tryptophan (W26) of the major coat protein, with probably no significant change in the alpha-helix content. Further contraction of the rodlike I-forms to spherical S-forms at 25 degrees C is accompanied by a substantial general decrease in the magnitude of the ellipticity throughout the 230-210-nm region, which is indicative of a decrease in the alpha-helix content of the major coat protein. The similarity of the circular dichroism spectrum of S-forms with that of coat protein in detergents suggests that the S-form coat protein resembles the coat protein in lipid bilayers. The intrinsic fluorescence of W26 is quenched without red-shift (but perhaps a barely detectable blue-shift) following fd contraction to I-forms and S-forms. The accessibility of W26 to aqueous quenchers does not change significantly upon contraction. However, interaction with hydrophobic quenchers is dramatically altered in the contracted forms in a manner suggesting that the environment surrounding the tryptophan changes from native-protein-like in the fd filament to molten globule-like in the I-form rods and S-form spheroids. As discussed herein, certain features of these data support previous suggestions that chloroform-induced filamentous phage contraction may provide information about phage penetration and assembly in vivo.