BINDING OF 2',3'-CYCLIC NUCLEOTIDE 3'-PHOSPHODIESTERASE TO MYELIN - AN IN-VITRO STUDY

The binding of 2',3'-cyclic nucleotide 3'-phosphodiesterase isoform 1 (CNP1) to myelin and its association with cytoskeletal elements of the sheath have been characterized with in vitro synthesized polypeptides and purified myelin. We have previously shown that the cysteine resid ue present in the carboxy-terminal CXXX box of CNP1 is isoprenylated, and that both C-15 farnesyl and C-20 geranylgeranyl isoprenoids can se rve as substrates for the modification. Here, we have mutated the CXXX box to obtain selectively farnesylated CNP1 or geranylgeranylated CNP 1 and found that these two modified forms of CNP1 behave identically i n all of the assays performed. Isoprenylation is essential but not suf ficient for the binding of in vitro synthesized CNP1 to purified myeli n, because a control nonmyelin protein is isoprenylated, yet unable to bind to myelin. In our assay, membrane-bound CNP1 partitions quantita tively into the nonionic detergent-insoluble phase of myelin, suggesti ng that CNP1 binds to cytoskeletal elements within myelin. However, is oprenylated CNP1 fails to bind to the cytoskeletal matrix isolated fro m myelin by detergent treatment, implying that both detergent-soluble and insoluble myelin components are involved in the binding of CNP1. A model for the interactions between CNP1 and myelin is presented, cons istent with models proposed for other isoprenylated proteins.