Human C4-binding protein. I. Isolation and characterization.
Human C4-binding protein. I. Isolation and characterization.
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DOI：
10.1084/jem.148.1.207
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发表时间：
1978-07-01
期刊：
影响因子：
--
通讯作者：
Nussenzweig V
中科院分区：
文献类型：
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作者：
Scharfstein J;Ferreira A;Gigli I;Nussenzweig V
C4结合蛋白（C4-bp）是补体系统的一种新成分，通过聚乙二醇沉淀，然后通过离子交换层析从人血浆中分离得到。在十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）上通过两个独立的标准鉴定C4-bp：其与C4 b结合的能力和与单特异性抗血清的免疫沉淀。纯化的C4-bp是一个10.7s的糖蛋白。它由几个分子量为70，000道尔顿的二硫键亚基组成。在非还原条件下，其分子量在SDS-PAGE上估计为540- 590，000道尔顿。C4-bp在pH8.6，不存在游离二价阳离子时作为缓慢的B-球蛋白移动，但当缓冲液含有Ca（++）-乳酸盐时，C4-bp是γ球蛋白。纯化的C4-bp与纯化的C4 b结合。该反应在二价阳离子存在或不存在下进行，并且不被二异丙基氟磷酸盐抑制。C4 b/C4-bp复合物在蔗糖梯度超离心上的沉降系数在15和17 s之间，并且可以容易地通过交叉免疫电泳（CIE）鉴定。复合物向阳极移动的速度比任何一种蛋白质都快。C4-bp是多价的。在C4 b/C4- bp的分子比为4至5时达到饱和。如果补体系统在运行前或运行期间被激活，C4 b和C4-bp之间的相互作用可能使正常人血清中这些蛋白质的电泳图谱复杂化。然而，在EDTA-血浆中，天然C4和C4-bp不形成稳定的复合物，并且可以在CIE之后在单独的峰中鉴定。
C4-binding protein (C4-bp), a new component of the complement system, was isolated from human plasma by precipitation with polyethyleneglycol, followed by chromatography on ion exchangers. C4-bp was identified on sodium dodecyl- sulfate polyacrylamide gel electrophoresis (SDS-PAGE) by two independent criteria: its ability to bind to C4b, and immunoprecipitation with a monospecificantiserum. Purified C4-bp is a 10.7 s glycoprotein. It consists of several disulfide bonded subunits of mol wt 70,000 daltons. Under nonreducing conditions, its mol wt has been estimated on SDS-PAGE as 540- 590,000 daltons. C4-bp moves as a slow B-globulin at pH 8.6 in the absence of free divalent cations, but when the buffers contain Ca(++)-lactate, C4-bp is a gamma globulin. Purified C4-bp binds to purified C4b. The reaction proceeds in the presence or absence of divalent cations and is not inhibited by diisopropylfluorophosphate. The C4b/C4-bp complexes have sedimentation coefficients between 15 and 17 s on sucrose gradient ultracentrifugation, and can be readily identified by crossed immunoelectrophoresis (CIE). The complexes move faster toward the anode than either protein. C4-bp is multivalent. Saturation is reached at molecular ratios of C4b/C4- bp of between 4 and 5. The interaction between C4b and C4-bp may complicate the electrophoretic patterns of these proteins in normal human serum, if the complement system is activated before or during the run. However, in EDTA-plasma, native C4 and C4-bp do not form stable complexes and can be identified in separate peaks after CIE.