Myelin proteolipid protein contains thioester-linked fatty acids.
Myelin proteolipid protein (PLP) is known to contain long-chain, covalently bound fatty acids. Previous studies, including our own, have suggested the occurrence of an oxyester type of linkage between fatty acids and PLP. However, we found that protein-SH groups are required in the acylation reaction, suggesting the possible presence of thioesters. In the present study, we have examined the nature of the acyl-PLP linkages by determining whether free thiol groups are generated on removal of fatty acids. Incubation of reduced and carboxyamidomethylated proteolipid apoprotein (RCM-APL) with 0.2 M hydroxylamine and [14C]iodoacetamide at pH 7.5 and 37 degrees C resulted in the release of fatty acids and the concomitant labeling of newly formed thiol groups. Incubation with Tris or methylamine at pH 7.5 failed to remove fatty acids and generate free -SH groups. The possibility that on treatment buried thiol groups became exposed was essentially excluded because (1) similar results were obtained in 2-chloroethanol, a solvent in which acylated and deacylated PLP have the same conformation, and (2) small PLP peptides were labeled only in the presence of hydroxylamine. On incubation with [14C]methylamine at pH 9.0, RCM-APL was not labeled, thus excluding the occurrence of intramolecular thiol esters. On the other hand, fatty acids were released as radioactive N-methyl fatty acylamide, indicating the presence of intermolecular thioesters between fatty acids and protein. These results demonstrate that a large proportion of fatty acids covalently bound to PLP are liked to -SH groups.
Bizzozero
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Biochemistry Department, E. K. Shriver Center, Waltham, MA 02254.
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D000215:Acylation; D000818:Animals; D002417:Cattle; D004952:Esters / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism*; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D007460:Iodoacetamide / Q000494:pharmacology; D008744:Methylamines / Q000494:pharmacology; D009185:Myelin Proteins / Q000378:metabolism*; D018991:Myelin Proteolipid Protein; D013438:Sulfhydryl Compounds / Q000378:metabolism*
D004952:Esters; D005227:Fatty Acids; D006898:Hydroxylamines; D008744:Methylamines; D009185:Myelin Proteins; D018991:Myelin Proteolipid Protein; D013438:Sulfhydryl Compounds; D019811:Hydroxylamine; C027451:methylamine; D007460:Iodoacetamide