Higgins J A
Biochim Biophys Acta. 1981 Jan 8;640(1):1-15. doi: 10.1016/0005-2736(81)90527-7.
Phosphatidylethanolamine of rat liver microsomes is rapidly methylated by S-adenosyl[methyl-14C]methionine to produce phosphatidyl-N-monomethylethanolamine, phosphatidyl-N,N-dimethylethanolamine and phosphatidylcholine. Using phospholipase C as a probe, on both opened (0.4% taurocholate or French pressure cell treatment) and unopened microsomes, it is demonstrated that phosphatidylcholine is labelled in the inner leaflet of the bilayer and, to a greater extent, in the outer leaflet. Phosphatidyl-N,N-dimethylethanolamine is labelled in the outer leaflet and in a pool sequestered from phospholipase C in open and closed vesicles. Phosphatidyl-N-monomethylethanolamine is labelled in a similarly sequestered pool. When microsomes containing labelled phosphatidyl-N-monomethylethanolamine and phosphatidyl-N,N-dimethylethanolamine were incubated with unlabeled S-adenosylmethionine, these phospholipids were methylated to produce phosphatidylcholine in the outer leaflet. This metabolism was inhibited by S-adenosylhomocysteine. Trypsin treatment of unopened microsomes inhibited 95% of the incorporation of 14CH3 into the outer leaflet of the bilayer with no effect on incorporation into sequestered phosphatidyl-N-monomethylethanolamine, phosphatidyl-N,N-dimethylethanolamine and phosphatidylcholine. Therefore, sequestered phosphatidyl-N-monomethylethanolamine and phosphatidyl-N,N-dimethylethanolamine are apparently synthesized by enzymes located at the inner surface of the microsomal membranes. These observations suggest that initial methylation of phosphatidylethanolamine takes place at the inner surface of the microsomes and that phosphatidyl-N-monomethylethanolamine is transferred to the outer leaflet to produce phosphatidylcholine. However, phosphatidyl-N-monomethylethanolamine is also methylated at the inner leaflet to produce phosphatidylcholine which does not equilibrate with that of the outer leaflet. Phosphatidylcholine of both the inner and outer bilayer leaflets is uniformly labelled by injection of [14C]methionine, in vivo.
大鼠肝微粒体的磷脂酰乙醇胺可被S-腺苷基[甲基-14C]甲硫氨酸迅速甲基化，生成磷脂酰-N-单甲基乙醇胺、磷脂酰-N,N-二甲基乙醇胺和磷脂酰胆碱。使用磷脂酶C作为探针，在经开放处理（0.4%牛磺胆酸盐或法国压榨细胞处理）和未经开放处理的微粒体上均证明，磷脂酰胆碱在双层膜的内小叶被标记，且在外小叶的标记程度更高。磷脂酰-N,N-二甲基乙醇胺在外小叶以及开放和封闭囊泡中与磷脂酶C隔离的一个库中被标记。磷脂酰-N-单甲基乙醇胺在类似的隔离库中被标记。当含有标记的磷脂酰-N-单甲基乙醇胺和磷脂酰-N,N-二甲基乙醇胺的微粒体与未标记的S-腺苷甲硫氨酸一起孵育时，这些磷脂会甲基化，在外小叶生成磷脂酰胆碱。这种代谢受到S-腺苷同型半胱氨酸的抑制。用胰蛋白酶处理未经开放处理的微粒体可抑制95%的14CH3掺入双层膜的外小叶，而对掺入隔离的磷脂酰-N-单甲基乙醇胺、磷脂酰-N,N-二甲基乙醇胺和磷脂酰胆碱没有影响。因此，隔离的磷脂酰-N-单甲基乙醇胺和磷脂酰-N,N-二甲基乙醇胺显然是由位于微粒体膜内表面的酶合成的。这些观察结果表明，磷脂酰乙醇胺的初始甲基化发生在微粒体的内表面，且磷脂酰-N-单甲基乙醇胺被转移到外小叶以生成磷脂酰胆碱。然而，磷脂酰-N-单甲基乙醇胺也在内小叶甲基化生成磷脂酰胆碱，且其与外小叶中的磷脂酰胆碱不会达到平衡。通过在体内注射[14C]甲硫氨酸，双层膜内小叶和外小叶的磷脂酰胆碱均被均匀标记。
