The amino acid sequence of the α-chain of human fibrinogen

Doolittle, Russell F.; Watt, K W; Cottrell, Barbara A.; Strong, Donna D.; Riley, Marcia

doi:10.1038/280464a0

Cited by 321 publications

(180 citation statements)

References 26 publications

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“…The minor band at ca. 2,500 nucleotides, rather than being another Aa mRNA species, probably represents cross-reaction of the Aa probe with BP mRNA, since it has been shown in mammals that these two subunits share some sequence homology (11). This conclusion is supported by the fact that this cross-reacting RNA was distributed into precisely the same RNA fractions which hybridized with the rat BP clone (compare Fig.…”

supporting

confidence: 70%

Xenopus fibrinogen: characterization of the mRNAs for the three subunits.

Holland¹,

Wangh²

1984

Mol. Cell. Biol.

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We purified and characterized the mRNAs coding for each of the three subunits of Xenopus fibrinogen. Purification was accomplished by electrophoretic separation of liver polyadenylated RNA in a fully denaturing gel, followed by recovery of the RNA from the gel via transfer to an ion-exchange membrane. This procedure yielded fractions which were highly enriched for the mRNAs for each of the fibrinogen chains. The fibrinogen mRNAs were identified by two methods: (i) in vitro translation followed by subunit-specific cleavage with the proteases thrombin and batroxobin; and (ii) cross-hybridization with cDNA clones for individual subunits of rat fibrinogen. The results demonstrate that the Aca and -y chains of frog fibrinogen are each coded by a single mRNA species. The Aca mRNA is ca. 3,100 nucleotides in length, which is nearly twice the minimum size required to code for the Aca precursor polypeptide. The y chain mRNA comprises about 1,600 bases and includes only a small untranslated region. In contrast, the BP subunit is synthesized from two mRNAs, one of which is 2,500 and the other 1,800 nucleotides long. The 2,500-base mRNA includes a large noncoding region, whereas the smaller one is near the minimum required size. The larger BP8 mRNA is ca. fivefold more abundant that the smaller species.Fibrinogen, which is synthesized and secreted by the liver, is found in the plasma of all vertebrates. The intact molecule comprises two sets of three nonidentical subunits designated Aax, BP, and -y. During the process of blood coagulation, the proteolytic enzyme thrombin cleaves the NH2-terminal A and B fibrinopeptides from the Aa and BP chains, thus generating fibrin which forms the structural matrix of a blood clot.We have previously described the polypeptide chain composition of fibrinogen from the frog Xenopus laevis (14,33 inducing the acute-phase response in mammals (16,19), was used in an effort to increase fibrinogen production in frogs. Polyadenylated RNA was purified (13) from the liver offrogs that had received a single injection of pure spirits of turpentine (Parks) into the leg muscle at a dose of 83 ,Il/50 g of body weight 12 to 24 h before sacrifice. This mRNA preparation was slightly enriched for the fibrinogen mRNAs relative to albumin mRNA, based on the in vitro translation assay, and was therefore used for the experiments described here. The sizes of all of the fibrinogen translation products and mRNAs were identical whether the RNA had been isolated from normal or turpentine-treated animals.To isolate the mRNAs coding for the Aa, BP, and -y chains of fibrinogen, liver mRNA was fractionated in a methylmercuric hydroxide agarose gel and recovered from the gel by transfer to and elution from a DEAE membrane (13). The high resolution of the RNA achieved during electrophoresis was preserved by slicing the DEAE membrane into 1-mm sections. The RNA eluted from each membrane slice was translated in vitro, and the products were resolved in a sodium dodecyl sulfate (SDS)-polyacrylamide gel (Fig. 1). All translated...

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supporting

confidence: 70%

Xenopus fibrinogen: characterization of the mRNAs for the three subunits.

Holland¹,

Wangh²

1984

Mol. Cell. Biol.

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“…Plasmin also cleaves fibrinogen (34) which, like fibronectin, contains the cytoadhesive tetrapeptide, Arg-Gly-Asp-Ser (35). Neutrophils resuspended in afibrinogenemic plasma released levels of elastase similar to that of normal plasma.…”

Section: Discussionmentioning

confidence: 99%

Fibronectin degradation products containing the cytoadhesive tetrapeptide stimulate human neutrophil degranulation.

Wachtfogel

Abrams²,

Kucich³

et al. 1988

J. Clin. Invest.

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We investigated whether adhesive glycoproteins, such as fibronectin or fibrinogen, could function to provide a nidus for neutrophil degranulation. Elastase release in recalcified plasma was normal in afibrinogenemic plasma, but 73% less in plasma depleted of fibronectin. Proteolytic digests of fibronectin, but not intact fibronectin (50-1,000 gg/ml), induced a concentration-dependent release of neutrophil elastase and lactoferrin. MAbs N293, which recognized the mid-molecule of fibronectin, N294, which was directed toward the 11-kD cell adhesive fragment, and N295, generated against the amino terminal of the 11-kD fragment, inhibited the release of elastase by 7, 24, and 60%, respectively. The cytoadhesive tetrapeptide portion of fibronectin, Arg-Gly-Asp-Ser (250-1,000 pg/ml), released 1.94±0.10 jig/ml of elastase from 107 neutrophils, in contrast to the lack of release by the control hexapeptide, Arg-GlyTyr-Ser-Leu-Gly. Plasmin appeared to be the enzyme responsible for fibronectin cleavage, since neutrophil elastase release in plasma that had been depleted of plasminogen was decreased and reconstitution of plasminogen-deficient plasma with purified plasminogen corrected the abnormal release. Plasmin cleaved fibronectin to multiple degradation products, each < 200 kD. This fibronectin digest released 1.05 jig/ml of elastase from 107 neutrophils. We suggest that the activation of plasminogen leads to the formation of fibronectin degradation products capable of functioning as agonists for neutrophils.

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“…By far the most unstructured of these is the C-terminal end of the ␣-subunits termed ␣CA and ␣CB. In the crystal structure of chicken fibrinogen (1), ␣CA and ␣CB appeared disordered in the electron density map, despite the fact that the chicken protein is shorter by 119 residues and lacks the 10 ϫ 13 repeated sequences (24,25) that add additional floppiness to the bovine and human proteins. These ␣C domains termed ''free-swimming appendages'' often interfere with crystal formation and had to be proteolyzed in the structure published by Brown et al (2).…”

Section: Discussionmentioning

confidence: 99%

Evidence for S -nitrosothiol-dependent changes in fibrinogen that do not involve transnitrosation or thiolation

Akhter

Vignini

Zhang

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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S-nitrosoglutathione (GSNO, 50 M) inhibited the initial rate of thrombin-catalyzed human and bovine fibrinogen polymerization by Ϸ50% to 68% respectively. Inhibition was also observed with other structurally varied S-nitrosothiols (RSNOs) including sugar derivatives of S-nitroso-N-acetylpenicillamine (SNAP). The fact that the same concentration of GSNO had no effect on thrombindependent hydrolysis of tosylglycylprolylarginine-4-nitroanilide acetate suggested that this inhibition was due to GSNO-induced changes in fibrinogen structure. This result was confirmed by CD spectroscopy where GSNO or S-nitrosohomocysteine increased the ␣-helical content of fibrinogen by Ϸ15% and 11%, respectively. S-carboxymethylamido derivatives of glutathione or homocysteine had no effect on the fibrinogen secondary structure. The GSNO-dependent secondary structural effects were reversed on gel filtration chromatography, suggesting that the effects were allosteric. Further evidence for fibrinogen-GSNO interactions was obtained from GSNO-dependent quenching of the intrinsic fibrinogen Trp fluorescence and the perturbation of the GSNO circular dichroic absorbance as a function of [fibrinogen]. The K ds of 3 to 10 M for fibrinogen-GSNO interactions with a stoichiometry of 2:1 (GSNO:fibrinogen) were estimated from isothermal titration calorimetry and fluorescence quenching, respectively. These results suggest that RSNOs induce changes to fibrinogen structure by interacting at specific aromatic rich domains. Three such putative RSNO-binding domains have been identified in the unordered, aromatic residue-rich C-termini of the ␣-chains of fibrinogen.

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The amino acid sequence of the α-chain of human fibrinogen

Cited by 321 publications

References 26 publications

Xenopus fibrinogen: characterization of the mRNAs for the three subunits.

Xenopus fibrinogen: characterization of the mRNAs for the three subunits.

Fibronectin degradation products containing the cytoadhesive tetrapeptide stimulate human neutrophil degranulation.

Evidence for S -nitrosothiol-dependent changes in fibrinogen that do not involve transnitrosation or thiolation

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