THE PRESENCE OF UBIQUITIN-PROTEIN CONJUGATES IN PLANT CHLOROPLAST LYSATES IS DUE TO CYTOSOLIC CONTAMINATION

Chloroplasts were isolated by Percoll density gradient centrifugation from Lemna minor and Spinacia oleracea grown under normal conditions o r from L. minor subjected to sulfur starvation, a condition that induc es a dramatic increase in the rates of proteolysis, namely in the degr adation of the chloroplast enzyme ribulosebisphosphate carboxylase. Th e presence of free ubiquitin or ubiquitin-protein conjugates was not i mmunologically detected in the purified chloroplast extracts under eit her of these conditions. Moreover, no ATP-dependent conjugation of I-1 25-labelled ubiquitin to endogenous proteins was detected in L. minor chloroplast lysates. Our results support the view that plant chloropla sts do not possess free ubiquitin or ubiquitin conjugating activity. F urthermore, treatment of the mechanically isolated chloroplasts with t hermolysin suggests that the presence of ubiquitin-protein conjugates in some chloroplast preparations is due to cytosolic contamination.