Polarized Fourier transform infrared (FTIR) difference spectroscopy of the M412 intermediate in the bacteriorhodopsin photocycle

The possibility that light-induced protein conformational changes accompany the formation of the M₄₁₂ species in the bacteriorhodopsin photocycle is investigated by polarized Fourier transform infrared (FTIR) spectroscopy on oriented films of purple membrane. From the light-induced FTIR dichroism changes, it is estimated that: (i) the CO stretching vibration at 1762 cm⁻¹, which has been assigned to a protonated Asp carboxyl group in M₄₁₂ [(1985) Biochemistry 24, 400-407], is oriented at (θ = 35 ± 5° from the normal to the membrane plane; (ii) the limit for the change in the average tilt angle of the α-helices after photoconversion is less than 2°. The latter observation excludes the large variations in the protein conformation during the M⁴¹² formation proposed by Draheim and Cassim [(1985) Biophys. J. 47, 497-507].