Plasma and Platelet Fibrinogen Differ in γ Chain Content

Charles W Francis; Ralph L Nachman; Victor J Marder

doi:10.1055/s-0038-1661026

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1984; 51(01): 084-088
DOI: 10.1055/s-0038-1661026

Original Article

Schattauer GmbH Stuttgart

Plasma and Platelet Fibrinogen Differ in γ Chain Content

Authors

Charles W Francis

The Hematology Unit, Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York, N. Y., U.S.A.
Ralph L Nachman

¹The Cornell University Medical Center, New York, N. Y., U.S.A.
Victor J Marder

The Hematology Unit, Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York, N. Y., U.S.A.

Abstract

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Summary

The polypeptide chain composition of fibrinogen and cross- linked fibrin from normal platelets and plasma has been compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Fibrinogen was prepared from a lysate of normal human platelets by ethanol precipitation and by antifibrinogen immunoaffinity chromatography. While the Aα chains of platelet fibrinogen appeared to be somewhat degraded, the electrophoretic mobilities of purified platelet and plasma fibrinogen Bβ an γ chains were the same. Crosslinked fibrin was prepared from platelet and plasma fibrinogen and γ chain dimers identified by their electrophoretic mobility, incorporation of the lysine analog dansyl cadaverine during crosslinking, and by reaction with antifibrinogen antibody in Western blots. Platelet crosslinked fibrin had a different polypeptide chain composition than that of plasma crosslinked fibrin with absence of the γ₅₀–γ_57.5 dimer in the platelet fibrin. This finding indicates that the γ_57.5 chain, which constitutes 5% of total normal plasma fibrinogen γ chains, is either absent or present in markedly reduced amounts in platelet fibrinogen.

Key Words

Fibrinogen - Fibrin - Platelets - γ Chain

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References

References
1 Ware AG, Fahey JL, Seegers WH. Platelet extracts, fibrin formation and interaction of purified prothrombin and thromboplastin. Am J Physiol 1948; 154: 140-147
2 Grette K. Extraction and partial purification of platelet-fibrinogen. In: Studies on the Mechanism of Thrombin-Catalyzed Hemostatic Reactions in Blood Platelets. Acta Physiol Scand 1962; Suppl 195: 32-36
3 Gokcen M, Yunis E. Fibrinogen as a part of platelet structure. Nature 1963; 200: 590-591
4 Bezkorovainy A, Rafelson ME. Characterization of some proteins from normal human platelets. J Lab Clin Med 1964; 64: 212-225
5 Castaldi PA, Caen J. Platelet fibrinogen. J Clin Pathol 1965; 18: 579-585
6 Nachman RL. Immunologic studies of platelet protein. Blood 1965; 25: 703-711
7 Nachman RL, Marcus AJ, Zucker-Franklin D. Immunologic studies of proteins associated with subcellular fractions of normal human platelets. J Lab Clin Med 1967; 69: 651-658
8 Doolittle RF, Takagi T, Cottrell BA. Platelet and plasma fibrinogens are identifical gene products. Science 1974; 185: 368-370
9 Solum NO, Lopaciuk S. Bovine platelet proteins. III. Some properties of platelet fibrinogen. Thrombos Diathes Haemorrh 1969; 21: 428-440
10 Ganguly P. Isolation and some properties of fibrinogen from human blood platelets. J Biol Chem 1972; 247: 1809-1816
11 James HL, Bradford HR, Ganguly PPlatelet. Platelet fibrinogen. I. Identity and initial observations on the mode of its degradation by plasmin. Biochim Biophys Acta 1975; 386: 209-220
12 James HL, Ganguly P. Identity of human platelet fibrinogen. Biochem Biophys Res Comm 1975; 63: 659-662
13 Plow EF, Edgington TS. Unique immunochemical features and intracellular stability of platelet fibrinogen. Thromb Res 1975; 7: 729-742
14 James HL, Ganguly P, Jackson CW. Characterization and origin of fibrinogen in blood platelets. A review with recent data Thromb Haemostas 1977; 38: 939-954
15 Jandrot-Perrus M, Mosesson MW, Denniger M-H, Menache D. Studies of platelet fibrinogen from a subject with a congenital plasma fibrinogen abnormality (Fibrinogen Paris I). Blood 1977; 54: 1109-1116
16 Soria J, Soria C, Samama M, Poirot E, Kling C. Human platelet fibrinogen: A protein different from plasma fibrinogen. Pathol Biol 1976; 24 (Suppl) 15-17
17 Francis CW, Marder VJ, Martin SE. Demonstration of a large molecular weight variant of the γ chain of normal human plasma fibrinogen. J Biol Chem 1980; 255: 5599-5604
18 Wolfenstein-Todel C, Mosesson MW. Human plasma fibrinogen heterogeneity: Evidence for an extended carboxyl-terminal sequence in a normal γ chain variant (γ). Proc Natl Acad Sci 1980; 77: 5069-5973
19 Ferguson EW. High-resolution two-dimensional electrophoretic analysis of fibrinogen digestion by plasmin. J Lab Clin Med 1980; 96: 710-721
20 Francis CW, Kraus DH, Marder VJ. Structural and chromatographic heterogeneity of normal plasma fibrinogen associated with the presence of three y-chain types with distinct molecular weights. Biochim Biophys Acta 1983; 744: 155-164
21 Wolfenstein-Todel C, Mosesson MW. Carboxy-terminal amino acid sequence of a human fibrinogen γ-chain variant (γ). Biochemistry 1981; 20: 6146-6149
22 Crabtree GR, Kant JA. Organization of the rat γ-fibrinogen gene: Alternative mRNA splice patterns produce the γA and γB (γ) chains of fibrinogen. Cell 1982; 31: 159-166
23 Broekman MJ, Westmoreland NP, Cohen P. An improved method for isolating alpha granules and mitochondria from human platelets. J Cell Biol 1974; 60: 507-519
24 Doolittle RF. Fibrinogen. In: CRC Handbook Series in Clinical Laboratory Science, Section I: Hematology. Seligson D, Schmidt RM. (Eds.) 1980. 3 3-14
25 Cohn EJ, Strong LE, Hughes Jr WL, Mulford DJ, Ashworth JN, Melin M, Taylor HL. Preparation and properties of serum and plasma proteins. IV A system for the separation into fractions of the protein and lipoprotein component of biological tissues and fluids. J Am Chem Soc 1946; 68: 459-475
26 March SC, Parikh I, Cuatrecasas P. A simplified method for cyanogen bromide activation of agarose for affinity chromatography. Anal Biochem 1974; 60: 149-152
27 Porath J, Axen R, Emback S. Chemical coupling of proteins to agarose. Nature 1967; 215: 1491-1492
28 Ochterlony O. Diffusion-in-gel methods for immunological analysis. Prog Allergy 1958; 5: 1-78
29 Lorand L, Urayama T, de Kiewiet JW C, Nossel HL. Diagnostic and genetic studies on fibrin-stabilizing factor with a new assay based on amine incorporation. J Clin Invest 1969; 48: 1054-1064
30 Loewy AG, Dunathan K, Kriel R, Wolfinger Jr HL. Fibrinase. I. Purification of substrate and enzyme. J Biol Chem 1962; 236: 2625-2643
31 Lorand L, Chenoweth D, Gray A. Titration of the acceptor crosslinking sites in fibrin. Ann N Y Acad Sci 1972; 202: 155-171
32 Margolis J, Kenrick KG. Polyacrylamide gel-electrophoresis across a molecular sieve gradient. Nature 1967; 214: 1334-1336
33 Neville Jr DM. Molecular weight determination of protein-dodecyl sulfate complexes by gel electrophoresis in a discontinuous buffer system. J Biol Chem 1971; 246: 6328-6334
34 Francis CW, Marder VJ, Martin SE. Detection of circulating crosslinked fibrin derivatives by a heat extraction-SDS gradient gel electrophoretic technique. Blood 1979; 54: 1282-1295
35 Weber K, Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem 1969; 244: 4406-4412
36 Fairbanks G, Steck T, Walbach D. Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry 1971; 10: 2606-2617
37 Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications. Proc Natl Acad Sd 1969; 76: 4350-4354
38 Cooper IA, Firkin BG. Amino acid transport in human platelets and subsequent incorporation into protein. Thrombos Diathes Haemorrh 1970; 23: 140-147
39 Nachman R, Levine R, Jaffe E. Synthesis of actin by cultured guinea pig megakaryocytes. Complex formation with fibrin Biochim Biophys Acta 1978; 543: 91-105
40 Hawiger J, Timmons S, Kloczewiak M, Strong DD, Doolittle RF. γ and α chains of human fibrinogen possess sites reactive with human platelet receptors. Proc Natl Acad Sci 1982; 79: 2068-2071
41 Kloczewiak M, Timmons S, Hawiger J. Localization of a site interacting with human platelet receptor on carboxy-terminal segment of human fibrinogen γ chain. Biochem Biophys Res Comm 1982; 107: 181-187
42 Alt FW, Bothwell AL M, Knapp M, Siden E, Mather E, Koshland M, Baltimore D. Synthesis of secreted and membrane-bound immunoglobulin mu heavy chains is directed by mRNAs that differ at their 3 ends. Cell 1980; 20: 293-301
43 Early P, Rogers J, Davis M, Calame K, Bond M, Wall R, Hood L. Two mRNAs can be produced from a single immunoglobulin p gene by alternative RNA processing pathways. Cell 1980; 20: 313-319
44 Singer PA, Singer HH, Williamson AR. Different species of messenger RNA encode receptor and secretory IgM p chains differing at the carboxy termini. Nature 1980; 285: 294-300
45 Tyler BM, Cowman AF, Adams JM, Harris AW. Generation of long mRNA for membrane immunoglobulin γ2a chains by differential splicing. Nature 1981; 293: 406-408