Anticoagulant And Calcium-Binding Properties Of High Molecular Weight Derivatives Of Human Fibrinogen, Produced By Plasmin (Fragments X)

1981 ◽  
Author(s):  
W Nieuwenhuizen ◽  
M Gravesen
Keyword(s):  
Calcium Ions ◽  
Calcium Binding ◽  
Degradation Products ◽  
Amino Acid Sequences ◽  
Binding Properties ◽  
Human Fibrinogen ◽  
Molecular Weights ◽  
Amino Terminal ◽  
Terminal Amino ◽  
Carboxy Terminal

Early plasmin degradation products (fragments X) of human fibrinogen were prepared and purified on Sepharose 6B-CL. X-fragments were characterized with respect to amino-terminal amino acid sequences, polypeptide-chain composition, anticlotting properties and calcium-binding. Amino-terminal amino acids were alanine and tyrosine. The molecular weights of the chains were about 26,000, 58,000 and 48,000 for Aα-, Bβ- and γ-chains, respectively. Fragments X were about 6 times as potent in anticlotting behaviour as D-fragments prepared in the presence of calcium ions. Calcium-binding properties were similar to those of fibrinogen: No differences were observed between fragments X prepared in the presence of calcium ions and those prepared in the presence of EGTA. Results indicate that the carboxy-terminal parts of the Aα-chains of fibrinogen are not involved in calcium-binding and that differences in chain-remnants as observed in late plasmic degradation products (which depend on the presence of calcium ions or EGTA in the incubation medium) are introduced beyond the stage of fragment X-formation.

Factors Influencing The Structure Of Terminal Plasmin Degradation Products Of Human Fibrinogen And Fibrin

1981 ◽  
Author(s):  
W Nieuwenhuizen ◽  
A Vermond ◽  
F Haverkate
Keyword(s):  
Protective Effect ◽  
Calcium Ions ◽  
Degradation Products ◽  
Iminodiacetic Acid ◽  
Human Fibrinogen ◽  
Factors Influencing ◽  
Separate Effect ◽  
The Media ◽  
Carboxy Terminal ◽  
D Dimer

Experiments have been carried out with fibrinogen and with purified degradation products of fibrinogen and fibrin which demonstrate that the structure of D fragments obtained after prolonged plasmin digestion is influenced by several factors in the media.The previously described protective effect of calcium ions on the γ-chain carboxy-terminals of fibrinogen against plasmin attack is rather independent of the composition of the media (e.g., also observed in 2 M urea and/or high plasmin activities).Several compounds such as EDTA, EGTA, citrate and iminodiacetic acid appear to have a separate effect, which is best observed at low plasmin concentrations and in the absence of Ca2+ . Under these conditions, these compounds appear to make the γ-chain carboxy-terminal ends of the D- and D-dimer fragments more susceptible to plasmin digestion.Finally, as demonstrated by experiments with purified D;E complexes from fibrinogen and with whole fibrinogen digests, the E-moiety of the D:E complexes appears to be capable of protecting the D-moiety against low plasmin concentrations also in the absence of calcium ions.

THE USE OF REGION SPECIFIC RADIOIMMUNOASSAYS FOR CHARACTERIZATION OF CIRCULATING CALCITONIN IN PATIENTS WITH MEDULLARY CARCINOMA OF THE THYROID GLAND

1979 ◽  
Vol 91 (3) ◽  
pp. 449-461 ◽  
Author(s):  
Lisbeth Myhre ◽  
Kaare M. Gautvik
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Molecular Forms ◽  
Amino Acid Residues ◽  
Terminal Amino Acid ◽  
Amino Terminal ◽  
Terminal Fragment ◽  
Terminal Amino ◽  
Carboxy Terminal ◽  
Amino Terminal Fragment

ABSTRACT Two antisera with known region specificities have been used to characterize calcitonin immunoreactivity (iCT) in serum of patients with medullary thyroid carcinoma (MCT). Antiserum I which was raised against the synthetic hormone (1–32 amino acid residues), contained heterogeneous populations of immunoglobulins directed predominantly against carboxyterminal sequences of the hormone, but the antiserum reacted also with the amino-terminal fragment (1–10 amino acid residues). Antiserum II, which was raised against the carboxy-terminal hormone fragment (11–32 amino acid residues) reached equally well with the intact hormone and the C-terminal fragment, but showed negligible binding of the amino terminal fragment. Antiserum I measured therefore both amino-terminal and carboxy-terminal sequences of calcitonin while antiserum II measured only carboxy-terminal amino acid sequences. In 40 patients with MCT, antiserum I measured usually the highest concentration of serum iCT suggesting the presence of non-uniform hormone immunoreactivity. The different molecular forms of circulating iCT in 7 MCT patients were explored by using antiserum I after gel filtration on Sephadex G-100. The patients who were selected on basis of iCT measurement in serum using antiserum I and II, could be divided into 3 groups which showed characteristic iCT profiles. Group 1, in which antiserum II measured a higher concentration of serum iCT, contained predominantly (60–70 %) small fragments of calcitonin immunoreactivity. On the other hand, in the sera of group 3 in which antisera I measured an equal or the highest concentrations, the dominant form of the hormone consisted of molecular sequences equal to or larger than the intact hormone (90 %). In group 2, the two antisera measured an equal amount of serum iCT and molecular forms consisting mostly of larger hormone fragments dominated (50 %). All the patients were normocalcaemic in spite of frequently grossly elevated serum iCT, and 33 out of 36 patients had normal serum immunoreactive parathyroid hormone. In conclusion: 1. Serum iCT is heterogeneous and represents peptides of quite different molecular size with no or low biological activity. 2. Most of the serum calcitonin immunoreactivity consists of peptides with carboxy-terminal amino acid sequences. 3. Most, if not all, of the amino-terminal calcitonin immunoreactivity is due to monomeric and polymeric hormonal forms.

Degradation Pathway of Fibrinogen by Plasmin

1977 ◽  
Vol 38 (04) ◽  
pp. 0793-0800 ◽  
Author(s):  
Andrei Z. Budzynski ◽  
Victor J. Marder
Keyword(s):  
Amino Acids ◽  
Disulfide Bonds ◽  
Polypeptide Chain ◽  
Degradation Products ◽  
Degradation Pathway ◽  
Molecular Weights ◽  
Amino Terminal ◽  
Terminal Amino ◽  
Covalently Linked ◽  
Fibrinogen Molecule

SummaryThe molecular weights of derivatives obtained from chemical and enzymatic degradation of fibrinogen and fibrin support a model in which the two halves of the fibrinogen molecule are covalently linked by a set of disulfide bonds at the amino-terminal region. The 2 asymmetric cleavages caused by plasmin in the fibrinogen molecule occur according to the reactions:X → Y + DY → E + DThe quantitative analysis of the amino-terminal amino acids in fragments D (from fibrinogen) and DD (from crosslinked fibrin) yields a total of 3.0 and 6.9 moles of amino acids per mole of protein, indicating three and six polypeptide chain structures, respectively. The data on molecular weights, polypeptide chain composition and immunologic properties of fibrinogen degradation products support the hypothesis on the asymmetric pathway of fibrinogen degradation by plasmin and the formation of two fragment D and one fragment E molecules from each molecule of fibrinogen.

Permeability Enhancing and Chemotactic Activities of Lower Molecular Weight Degradation Products of Human Fibrinogen

1981 ◽  
Vol 45 (01) ◽  
pp. 090-094 ◽  
Author(s):  
Katsuo Sueishi ◽  
Shigeru Nanno ◽  
Kenzo Tanaka
Keyword(s):  
Molecular Weight ◽  
Degradation Products ◽  
Electron Microscopic Observation ◽  
Chemotactic Activity ◽  
Lower Molecular Weight ◽  
Electron Microscopic ◽  
Human Fibrinogen ◽  
Rabbit Skin ◽  
Molecular Weights ◽  
Active Fragments

SummaryFibrinogen degradation products were investigated for leukocyte chemotactic activity and for enhancement of vascular permeability. Both activities increased progressively with plasmin digestion of fibrinogen. Active fragments were partially purified from 24 hr-plasmin digests. Molecular weights of the permeability increasing and chemotactic activity fractions were 25,000-15,000 and 25,000 respectively. Both fractions had much higher activities than the fragment X, Y, D or E. Electron microscopic observation of the small blood vessels in rabbit skin correlated increased permeability with the formation of characteristic gaps between adjoining endothelial cells and their contraction.These findings suggest that lower molecular weight degradation products of fibrinogen may be influential in contributing to granulocytic infiltration and enhanced permeability in lesions characterized by deposits of fibrin and/or fibrinogen.

N-Terminal amino acid sequence of rat tonin: homology with serine proteases

1978 ◽  
Vol 56 (9) ◽  
pp. 920-925 ◽  
Author(s):  
N. G. Seidah ◽  
R. Routhier ◽  
M. Caron ◽  
M. Chrétien ◽  
S. Demassieux ◽  
...  
Keyword(s):  
Serine Proteases ◽  
Terminal Sequence ◽  
Renin Substrate ◽  
Amino Terminal ◽  
Single Polypeptide Chain ◽  
Serine Protease Family ◽  
Terminal Amino ◽  
Single Polypeptide ◽  
Carboxy Terminal ◽  
Amino Terminal Sequence

In this paper, we present the amino-terminal sequence of rat tonin, an endopeptidase responsible for the conversion of angiotensinogen, the tetradecapeptide renin substrate, or angiotensin I to angiotensin II. It is shown that isoleucine and proline occupy the amino- and carboxy-terminal residues respectively. The N-terminal sequence analysis permitted the identification of 34 out of the first 40 residue s of the single polypeptide chain composed of 272 amino acids. The se results showed an extensive homology with the sequence of many serine proteases of the trypsin–chymotrypsin family. This information, coupled with the slow inhibition of tonin by diisopropylfluorophosphate, classified this enzyme as a selective endopeptidase of the active serine protease family.

Neisseria pili proteins: amino-terminal amino acid sequences and identification of an unusual amino acid

Biochemistry ◽  
1978 ◽  
Vol 17 (3) ◽  
pp. 442-445 ◽  
Author(s):  
Mark A. Hermodson ◽  
Kirk C. S. Chen ◽  
Thomas M. Buchanan
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Terminal Amino Acid ◽  
Amino Terminal ◽  
Terminal Amino ◽  
Unusual Amino Acid

Human Fibrinogen Binds Edta And Citrate

1981 ◽  
Author(s):  
W Nieuwenhuizen ◽  
A Vermond ◽  
J Hermans
Keyword(s):  
Calcium Ions ◽  
Equilibrium Dialysis ◽  
Strong Dependence ◽  
Direct Interaction ◽  
Heat Stability ◽  
Binding Studies ◽  
Human Fibrinogen ◽  
Direct Binding ◽  
Carboxy Terminal ◽  
Fibrinogen Molecule

It was shown that EDTA and citrate make the carboxy-terminal parts of the γ-chains of fibrinogen more susceptible to plasmin degradation. This is an effect independent of the calcium-chelating properties of those compounds. Furthermore, EDTA prolongs the thrombin times decreases the heat stability and causes the formation of abnormal clots.This suggests a direct interaction of EDTA (and possibly also of citrate) with the fibrinogen molecule thereby causing a conformational change. The interaction of citrate and EDTA with fibrinogen was measured by direct binding studies (equilibrium dialysis).It was found that at pH 7.5 human fibrinogen binds 3.4 molecules of citrate with Kd = 1.4 x 10-4M or 0.4 molecules of EDTA with Kd = 2.2 x 10-5M. Citrate and EDTA compete for the same binding site(s). No binding of acetate could be demonstrated.The binding of EDTA and citrate shows a strong ^-dependence suggesting a (partly) ionogenic binding between charged areas on the fibrinogen molecule and charged groups on EDTA or citrateOur results support the idea that the binding of EDTA and citrate and the concomitant effect on the plasmin attack are independent of the effect of calcium ions

Recalculation of calcium-binding properties of human and rat fibrin(ogen) and their degradation products

1981 ◽  
Vol 22 (5-6) ◽  
pp. 653-657 ◽  
Author(s):  
Willem Nieuwenhuizen ◽  
Irina A.M. van Ruijven-Vermeer ◽  
Willem J. Nooijen ◽  
Anton Vermond ◽  
Frits Haverkate ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Degradation Products ◽  
Binding Properties

scholarly journals Effect of N- and C-Terminal Amino Acids on the Interfacial Binding Properties of Phospholipase D from Vibrio parahaemolyticus

2018 ◽  
Vol 19 (8) ◽  
pp. 2447
Author(s):  
Fanghua Wang ◽  
Ruixia Wei ◽  
Abdelkarim Abousalham ◽  
Wuchong Chen ◽  
Bo Yang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Secondary Structure ◽  
Phospholipase D ◽  
Vibrio Parahaemolyticus ◽  
Terminal Segment ◽  
Amino Acid Sequences ◽  
Loop Structure ◽  
Binding Properties ◽  
Phospholipid Monolayers ◽  
Terminal Amino

The effects of N-terminal (1–34 amino acids) and C-terminal (434–487 amino acids) amino acid sequences on the interfacial binding properties of Phospholipase D from Vibrio parahaemolyticus (VpPLD) were characterized by using monomolecular film technology. Online tools allowed the prediction of the secondary structure of the target N- and C-terminal VpPLD sequences. Various truncated forms of VpPLD with different N- or C-terminal deletions were designed, based on their secondary structure, and their membrane binding properties were examined. The analysis of the maximum insertion pressure (MIP) and synergy factor “a” indicated that the loop structure (1–25 amino acids) in the N-terminal segment of VpPLD had a positive effect on the binding of VpPLD to phospholipid monolayers, especially to 1,2-dimyristoyl-sn-glycero-3-phosphoserine and 1,2-dimyristoyl-sn-glycero-3-phosphocholine. The deletion affecting the N-terminus loop structure caused a significant decrease of the MIP and synergy factor a of the protein for these phospholipid monolayers. Conversely, the deletion of the helix structure (26–34 amino acids) basically had no influence on the binding of VpPLD to phospholipid monolayers. The deletion of the C-terminal amino acids 434–487 did not significantly change the binding selectivity of VpPLD for the various phospholipid monolayer tested here. However, a significant increase of the MIP value for all the phospholipid monolayers strongly indicated that the three-strand segment (434–469 amino acids) had a great negative effect on the interfacial binding to these phospholipid monolayers. The deletion of this peptide caused a significantly greater insertion of the protein into the phospholipid monolayers examined. The present study provides detailed information on the effect of the N- and C-terminal segments of VpPLD on the interfacial binding properties of the enzyme and improves our understanding of the interactions between this enzyme and cell membranes.

Two-Component Anti-Staphylococcus aureusLantibiotic Activity Produced by Staphylococcus aureusC55

1998 ◽  
Vol 64 (12) ◽  
pp. 4803-4808 ◽  
Author(s):  
Maduwe A. D. B. Navaratna ◽  
Hans-Georg Sahl ◽  
John R. Tagg
Keyword(s):  
Specific Activity ◽  
Micrococcus Luteus ◽  
Fold Increase ◽  
Reversed Phase ◽  
Amino Acid Sequences ◽  
Bacteriocin Activity ◽  
Terminal Amino Acid ◽  
Molecular Weights ◽  
Terminal Amino ◽  
Distinct Peptide

ABSTRACT Staphylococcus aureus C55 was shown to produce bacteriocin activity comprising three distinct peptide components, termed staphylococcins C55α, C55β, and C55γ. The three peptides were purified to homogeneity by a simple four-step purification procedure that consisted of ammonium sulfate precipitation followed by XAD-2 and reversed-phase (C8 and C18) chromatography. The yield following C8 chromatography was about 86%, with a more-than-300-fold increase in specific activity. When combined in approximately equimolar amounts, staphylococcins C55α and C55β acted synergistically to kill S. aureus or Micrococcus luteus but not S. epidermidis strains. The N-terminal amino acid sequences of all three peptides were obtained and staphylococcins C55α and C55β were shown to be lanthionine-containing (lantibiotic) molecules with molecular weights of 3,339 and 2,993, respectively. The C55γ peptide did not appear to be a lantibiotic, nor did it augment the inhibitory activities of staphylococcin C55α and/or C55β. Plasmids of 2.5 and 32.0 kb are present in strain C55, and following growth of this strain at elevated temperature (42°C), a large proportion of the progeny failed to produce strong bacteriocin activity and also lost the 32.0-kb plasmid. Protoplast transformation of these bacteria with purified 32-kb plasmid DNA regenerates the ability to produce the strong bacteriocin activity.

Sign in / Sign up

Export Citation Format

Share Document