A cyclophilin-regulated PP2A-like protein phosphatase in thylakoid membranes of plant chloroplasts

Dephosphorylation of central photosynthetic proteins regulates their turnov er in plant thylakoid membranes, A membrane protein phosphatase from spinac h thylakoids was purified 13000-fold using detergent-engaged FPLC, The puri fied enzyme exhibited characteristics typical of eukaryotic Ser/Thr phospha tases of the PP2A family in that it was inhibited by okadaic acid (IC50 = 0 .4 nM) and tautomycin (IC50 = 25 nM), irreversibly bound to microcystin-aga rose, and recognized by a polyclonal antibody raised against a recombinant catalytic subunit of human PP2A. Furthermore, the anti-PP2A antibody inhibi ted protein dephosphorylation in isolated thylakoids. The phosphatase copur ified with TLP40, a cyclophilin-like peptidyl-prolyl isomerase located in t he thylakoid lumen. TLP40 could be released from the phosphatase immobilize d on microcystin-agarose by high-salt treatment.: Binding of cyclosporin A (CsA) to TLP40 led to thylakoid phosphatase activation,while cyclophilin su bstrates, prolyl-containing oligopeptides, inhibited protein dephosphorylat ion. This dephosphorylation could be modulated by CsA or oligopeptides only after the thylakoids had been ruptured to expose the lumenal membrane surf ace where the TLP40 is located. Regulation of the PP2A-like phosphatase at the outer thylakoid surface is likely to operate via reversible binding of TLP40 to the inner membrane surface. This is a first example of transmembra ne regulation in which the activity of phosphatase is altered-by the bindin g of a cyclophilin to-a site other than the active one. We propose that sig naling from TLP40 to the protein phosphatase coordinates dephosphorylation and protein folding, two processes required for protein turnover during the repair of photoinhibited photosystem II reaction centers.