THE NATURE OF THE MULTICOMPONENT M-INTERMEDIATE FORMATION IN THE BACTERIORHODOPSIN PHOTOCYCLE .2. 3 COMPONENTS OF M-INTERMEDIATE FORMATION OF THE D96N BACTERIORHODOPSIN MUTANT

Kinetics of L intermediate decay and M intermediate formation for the D96N mutant at pH 7.0, 20 degrees C can be well fitted with a sum of t hree components with time constants tau(1) = 55 msec, t2 = 220 msec, a nd tau(3) = 1 msec. The M intermediate formation with a time constant of 55 msec correlates with the decay of L intermediate having differen ce absorbance maximum at 550 nm, while the M formation with a time con stant of 220 msec correlates with the decay of L intermediate having a maximum at 530 nm. The difference spectrum of L intermediate with dec ay time constant of 1 msec is well fitted by a sum of spectra correspo nding to L(1) and L(2) intermediates, and the third M has a difference maximum at 404 nm, whereas the first two M intermediates have maxima at 412 nm. These data support our scheme of two parallel pathways of M formation followed by irreversible transition into M'.