Williams S E, Inoue I, Tran H, Fry G L, Pladet M W, Iverius P H, Lalouel J M, Chappell D A, Strickland D K
Biochemistry Laboratory, American Red Cross, Rockville, Maryland 20855.
J Biol Chem. 1994 Mar 25;269(12):8653-8.
Lipoprotein lipase (LPL) binds with high affinity to the low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP) and promotes binding, uptake, and degradation of normal triglyceride-rich lipoproteins in a process mediated by LRP (Chappell, D. A., Fry, G. L., Naknitx, M.A., Muhonen, L. E., Pladet, M. W., Iverius, P-H., and Strickland, D. K. (1993) J. Biol. Chem. 268, 14168-14175). To localize the portion of LPL that is responsible for interacting with LRP, fragments of LPL were expressed in bacteria. A fragment of human LPL containing the COOH-terminal domain (residues 313-448, designated LPLC) which lacks the catalytic site was able to bind to LRP. Purified LRP bound specifically to microtiter wells coated with LPL or LPLC with KD values of 2.8 and 5 nM, respectively. The effects of several mutations of LPLC were tested. Mutation of Lys407 to Ala reduced the affinity of LPLC for LRP by approximately 10-fold. Like native LPL, LPLC prevented the binding of activated alpha 2-macroglobulin and the 39-kDa receptor-associated protein to LRP and inhibited the internalization and degradation of activated alpha 2-macroglobulin and receptor-associated protein in cultured fibroblasts. LPLC also bound to 125I-labeled human normal triglyceride-rich lipoproteins and promoted their binding to purified LRP and to cultured cells. Mutation of Trp393 and Trp394 to Ala completely abolished the ability of LPLC to bind to lipoproteins, but had little effect on its interaction with LRP. These data indicate that the COOH-terminal domain of LPL may function both in binding lipoproteins and mediating their interaction with LRP.
脂蛋白脂肪酶（LPL）以高亲和力与低密度脂蛋白受体相关蛋白/α2-巨球蛋白受体（LRP）结合，并在由LRP介导的过程中促进正常富含甘油三酯的脂蛋白的结合、摄取和降解（查佩尔，D.A.，弗莱，G.L.，纳克尼特克斯，M.A.，穆霍嫩，L.E.，普拉德特，M.W.，伊维留斯，P-H.，和斯特里克兰，D.K.（1993年）《生物化学杂志》268，14168 - 14175）。为了定位LPL中负责与LRP相互作用的部分，LPL的片段在细菌中表达。人LPL的一个包含COOH末端结构域（残基313 - 448，命名为LPLC）的片段，该片段缺乏催化位点，能够与LRP结合。纯化的LRP分别以2.8和5 nM的KD值特异性结合到包被有LPL或LPLC的微量滴定孔上。测试了LPLC的几个突变的影响。将赖氨酸407突变为丙氨酸使LPLC对LRP的亲和力降低了约10倍。与天然LPL一样，LPLC阻止活化的α2-巨球蛋白和39 kDa受体相关蛋白与LRP结合，并抑制培养的成纤维细胞中活化的α2-巨球蛋白和受体相关蛋白的内化和降解。LPLC还与125I标记的人正常富含甘油三酯的脂蛋白结合，并促进它们与纯化的LRP和培养细胞的结合。将色氨酸393和色氨酸394突变为丙氨酸完全消除了LPLC与脂蛋白结合的能力，但对其与LRP的相互作用影响很小。这些数据表明，LPL的COOH末端结构域可能在结合脂蛋白以及介导它们与LRP的相互作用中都发挥作用。
