Peptides fibrinopeptides

Substitution of particular amino acids in the fibrinopeptides and at thrombin s cleavage sites in recombinant fibrinogen has been used to understand thrombin specificity, the relative rates of fibrinopeptide cleavage, and effects on polymerization. If fibrinopeptide B is replaced with a fibrinopeptide-A-like peptide, fibrinopeptide release is similar to release from the Aa chain (Mullin et al., 2000). Thus, the ordered release of fibrinopeptides is dictated by the specificity of thrombin for its substrates. 

While Lorand (1952) found one peptide (fibrinopeptide), Bettelheim (1956) reported the presence of two peptides, A and B, peptide A traveling faster toward the anode than peptide B in a paper electrophoresis experiment at pH 4.15. Peptide A was found to contain N-terminal glutamic acid, the N-terminal residue missing from fibrinogen, while peptide B had no detectable N-terminal group. Lorand s fibrinopeptide is identical with peptide A. 

Thrombin [EC 3.4.21.5], also known as fibrinogenase, catalyzes the hydrolysis of peptide bonds, exhibiting preferential cleavage for the Arg—Gly peptide bond. The enzyme, a member of the peptidase family SI, activates fibrinogen to fibrin and releases fibrinopeptide A and B. Thrombin, formed from prothrombin, is more selective in peptide hydrolysis than trypsin or plasmin. 

After the early discovery of a tyrosine 0-sulfate residue in bovine fibrinopeptide B, 15 this posttranslational modification which occurs ubiquitously in proteins was also detected in a series of biologically active peptides such as the neurohormones of the gastrin/cholecysto-kinin (CCK) family of peptides, phyllokinin, Leu-enkephalin, and the thrombin inhibitor hirudin listed in Table 1. 

Fibrinogen is an elongated molecule with an (apy)2 structure.524,541,5413 Thrombin cleaves specific Arg-Gly bonds in the a and P chains releasing short (14- tol6-residue) "fibrinopeptides" from the N termini of the peptide chains. This leaves Gly-Pro-Arg "knobs" at... 

Peptide chimeras of this type have proven extremely valuable for the examination of an array of recognition events. In conjunction with molecular modeling, peptidomimetic substrates and inhibitors of human thrombin were designed and synthesized to evaluate our proposed structure for the thrombin-bound conformation of fibrinopeptide A (Scheme 2i)J%]... 

TV-terminal sections of these three chains, which are close in evolutionary terms. The a and 3 chains are terminated by a short peptide sequence called a fibrinopeptide that ordinarily prevents the spontaneous polymerisation of fibrinogen. 

Appropriate standard peptides for calibration with monoisotopic molecular masses covering, for example, the mass range 500-3000 Da (27). Glu-fibrinopeptide-B is also used as external calibrant for MS" experiments (28). 

Top) peptide ladder sequencing principle. Phenyl isothiocyanate (PITC) produces phenylthiohydantoin (PTH) of the terminal amino acid and a new peptide with one less amino acid. Phenyl isocyanate (PIC), in low quantity, produces N-terminal phenylcarba-mate (PC) from a small fraction of each peptide. (Bottom) example of sequencing of [Glu1]fibrinopeptide B. Reproduced (modified) from Chait B.T., Wang R., Beavis R.C. and Kent S.B.H., Science, 262, 89, 1993, with permission. 

Thus, characteristic changes can be revealed in the low molecular weight fraction of blood plasma, and these could serve as a new molecular marker of intoxication with RVX and other OPs. The peptides of rats and humans begin with Gly from the N-end, and the preceding truncated amino acid is a neutral and hydroxyl-bearing one (Thr in rat fibrinopeptide and Ser in human fibrinopeptide) (Table 7.5). 

TABLE 7.5. Fragments of fibrinopeptide A found in the peptide fraction of rat blood plasma in control and after subchronic exposure to RVX, and those found in human blood plasma in normal state and after in v/tro exposure to RVX. Phosphorylated peptide ADTGTTSEFIDEGAGIR (analogous to peptide ADSGEGDFLAEGGGVR found in human blood plasma) was not found in rat blood... 

Figure 3 A ladder sequence through 6 cycles of [Glul] fibrinopeptide B carried out on a total of 17.5 picomoles of starting peptide.

Figure 5 Quantitative derivatisation of 50 fmol [Glul] fibrinopeptide B with a C-5 quaternary linked tag. The derivatisation was performed on the peptide sample already spotted onto the target slide.

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