The 32 000-dalton Q(B)-protein of photosystem II (PS II) is rapidly damaged and removed from isolated pea thylakoids during incubation in the light resulting in a loss of photosynthetic electron flow through PS II. This in vitro photoinhibition is similar to that previously reported with intact Chlamydomonas cells. The damage occurs at a faster rate in vitro, however, due to the inability of isolated thylakoids to synthesize replacement Q(B)-protein. The removal of the damaged Q(B)-protein does not require any soluble components of the chloroplast stroma and is unaffected by the protease inhibitors phenyl-methylsulfonylfluoride or antipain. Unlike the effect of trypsin, no low mol. wt. membrane-bound or soluble fragments of the labelled Q(B)-protein could be identified either by autoradiography or immunologically using polyclonal antibodies specific for the Q(B)-protein. The lightinduced damage to the Q(B)-protein (indicated by a loss of Q(B) functional activity), preceded the removal of the protein from the membrane. We conclude that photodamage of the Q(B)-protein generates a conformational change which renders the protein susceptible to attack by a highly efficient, intrinsic membrane protease.