Immunochemical determination of conformational equilibria for fragments of the B beta chain of fibrinogen.
Immunochemical determination of conformational equilibria for fragments of the B beta chain of fibrinogen.
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纤维蛋白原 B β 链片段构象平衡的免疫化学测定。
DOI：
10.1021/bi00325a011
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发表时间：
1985
期刊：
影响因子：
2.9
通讯作者：
Scheraga,HA
中科院分区：
文献类型：
--
作者：
Nagy,JA;Meinwald,YC;Scheraga,HA
贝克化学实验室，康奈尔大学，伊萨卡，纽约14853接收于1983年12月16日;修订的Mandarin pt接收于1984年10月2日摘要：用免疫化学方法比较了完整纤维蛋白原分子的α链和含有凝血酶水解区的纤维蛋白原α链的各种片段的构象[Sachs，D. H、Schechter，A. N.，伊斯特莱克，A. & Anfmsen，C. B。等人（1972）Proc. Acad. Sci. USA 69，3790]。通过用天然牛纤维蛋白原免疫家兔诱导抗纤维蛋白原抗体。用免疫吸附法分离出对HSP 70片段20-28中天然抗原决定簇有特异性的抗体群。该制备用于确定该决定簇的非天然和天然构象的相互转化的平衡常数的值。在天然纤维蛋白原、二硫键结（DSK）、β-淀粉酶、β-淀粉酶片段16-28、β-淀粉酶片段20-28和纤维蛋白肽B（Fp B）中测定该决定簇的值。125 I-标记的纤维蛋白原（125 I-F）用于测定，通过测量125 I-F与研究中的纤维蛋白原衍生物之间结合纯化抗体的竞争。F抗原区、DSK和Ep抗原区在4 ℃时的α =无穷大，分别为（5.9±3.5）× 10 ~（-3）和（1.2±0.7）× 10~（-3）*。在4C下，对于FpB片段16-28、FpB片段20-28和FpB的Kmn值小于（6.0±3.9）× 10~ 7。这些测量结果表明，F中的抗原区域不受碘化的干扰，并且0.6%的DSK分子，0.1%的DSK分子和小于0.00006%的较小片段在水溶液中采用抗原决定簇内的天然构象。α的值比早先获得的CNBrAa的值大2个数量级[Nagy，J.A.，Meinwald，Y. C.的方法，& Scheraga，. A.（1982）Biochemistry 21，1794]，本研究中涉及的抗原决定簇位于牛纤维蛋白原β链的残基20-28（对应于人β链的残基13-21）中。免疫化学结果和早期报道的动力学数据[Hageman，T. C.的方法，& Scheraga，. A. 05.《生物化学与生物物理学文献》，1977年。179，506]，表明长程相互作用对于分别与抗体和凝血酶相互作用的纤维蛋白原区域中的天然结构的稳定是必需的。此外，所观察到的抗-Bcl 2（20-28）N抗体结合完整的天然牛F分子的能力支持了这样的结论，即Arg-Gly区域可接近天然牛和人纤维蛋白原的Aa和β链中的凝血酶催化水解[Martinelli，R.一、& Scheraga，. A.（1980）Biochemistry 19，2343; Hanna，L.美国，Scheraga，.一、弗朗西斯，C. W.，& Marder，V. J.（1984）Biochemistry 23，4681]，
Baker Laboratory of Chemistry, Cornell University, Ithaca, New York 14853 Received December 16, 1983; Revised Manuscript Received October 2, 1984 abstract: The conformations of the ß chain of the intact fibrinogen moleculeand of various fragments of the ß chain of fibrinogen that contain the region that is hydrolyzed by thrombin have been compared by an immunochemical method [Sachs, D. H., Schechter, A. N., Eastlake, A., & Anfmsen, C. B.(1972) Proc. Natl. Acad. Sci. USA 69, 3790]. Anti-fibrinogen antibodies were induced in rabbits by immunization with native bovine fibrinogen. An antibodypopulation specific for the native antigenic determinant within the ß fragment 20-28 was isolated by immunoadsorption. This preparation was used to determine the value of the equilibrium constant for the interconversion of the nonnative and native conformations of this determinant. Values of were measured for this determinant within native fibrinogen, the disulfide knot (DSK), ß, ß fragment 16-28, ß fragment 20-28, and fibrinopeptide B (FpB). 125I-Labeled fibrinogen (125I-F) was used in the determination of by measuring the competition between 125I-F and the fibrinogen derivatives under study for binding to the purified antibody. Forthe antigenic region in F, the DSK, and ß, the values of at 4 C were infinity,(5.9±3.5) X 10-3, and (1.2±0.7) X 10~ 3***, respectively. The values of Kmn {for ß fragment 16-28, ß fragment 20-28, and FpB at 4 C were less than (6.0±3.9) X 10~ 7. These measurements indicate that the antigenic region in F is unperturbed by iodination and that 0.6% of the DSK molecules, 0.1% of the ß molecules, and less than 0.00006% of the smaller fragments adopt the nativeconformation within the antigenic determinant in aqueous solution. The value for ß is 2 orders of magnitudegreater than that obtained earlier for CNBrAa [Nagy, J. A., Meinwald, Y. C., & Scheraga,. A.(1982) Biochemistry 21, 1794], The antigenic determinant involved in this study is localized among residues 20-28 of the ß chain of bovine fibrinogen (corresponding to residues 13-21 of the human ß chain). The immunochemical results, and kinetic data reported earlier [Hageman, T. C., & Scheraga,. A.(1977) Arch. Biochem. Biophys. 179, 506], suggest that long-range interactions are necessary for the stabilization of the native structure in the region of fibrinogen that interacts with the antibody and with thrombin, respectively. In addition, the observed ability of the anti-Bß (20-28) N antibodies to bind to intact native bovine F molecules supports the conclusion that the Arg-Gly region is accessible to thrombin-catalyzed hydrolysis in both the Aa and ß chains of native bovine and human fibrinogen [Martinelli, R. A., & Scheraga,. A.(1980) Biochemistry 19, 2343; Hanna, L. S., Scheraga,. A., Francis, C. W., & Marder, V. J.(1984) Biochemistry 23, 4681],
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DOI：
10.1200/jco.1984.2.11.1235
发表时间：
1984
期刊：
Journal of clinical oncology : official journal of the American Society of Clinical Oncology
影响因子：
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作者：
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DOI：
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发表时间：
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期刊：
Journal of immunology (Baltimore, Md. : 1950)
影响因子：
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DOI：
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作者：
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