IDENTIFICATION OF A PLASTID PROTEIN INVOLVED IN VESICLE FUSION AND ORMEMBRANE-PROTEIN TRANSLOCATION/

Structural evidence has accumulated suggesting that fusion and/or tran slocation factors are involved in plastid membrane biogenesis. To test this hypothesis, we have de, eloped an in vitro system in which the e xtent of fusion and/or translocation is monitored by the conversion of the xanthophyll epoxide (antheraxanthin) into the red ketocarotenoid (capsanthin), Only chromoplast membrane vesicles from red pepper fruit s (Capsicum annum) contain the required enzyme. Vesicles prepared from the mutant yellow cultivar are devoid of this enzyme and accumulate a ntheraxanthin. The fusion and/or translocation activity is characteriz ed by complementation due to the synthesis of capsanthin and the paral lel decrease of antheraxanthin when the two types of vesicles are incu bated together in the presence of plastid stroma, We show that the ext ent of conversion is dependent upon an ATP-requiring protein that is s ensitive to N-ethylmaleimide. Further purification and immunological a nalysis have revealed that the active factor, designated plastid fusio n and/or translocation factor (Pftf), resides in a protein of 72 kDa. cDNA cloning revealed that mature Pftf has significant homology to yea st and animal (NSF) or bacterial (Ftsh) proteins involved in vesicle f usion or membrane protein translocation.