CONFORMATIONAL EQUILIBRIA IN THE γ CHAIN COOH-TERMINUS OF HUMAN FIBRINOGEN

1987 ◽  
Author(s):  
C S Cierniewski ◽  
A Z Budzynski
Keyword(s):  
Binding Sites ◽  
Synthetic Peptide ◽  
Native Structure ◽  
Native Conformation ◽  
Human Fibrinogen ◽  
Long Range Interactions ◽  
Conformational Equilibria ◽  
Crosslinking Site ◽  
Antibody Population

Synthetic peptides and fragments cleaved from native fibrinogen are used in studies to localize binding sites for various ligands. We addressed the question how the native conformation of a selected γ chain segment is affected by scission of the original chain. The conformation of the γ chain COOH-terminus of intact fibrinogen and its various fragments containing this region has been compared by an immunochemical analysis. An antibody population specific for the native epitope within the γ391-405 segment was isolated by affinity chromatography on the corresponding synthetic peptide. Between 19.2 and 22.8% of antibodies were obtained from three different antisera indicating that this region represents one of the major epitopes of native fibrinogen. Anti-γ391-405 antibodies were used to determine the value of Kconf the equilibrium constant for the interconversion of the non-native and native conformations of this epitope. The measurements were done using native fibrinogen, fragments D1 and DD, γ chain and γ391-405 synthetic peptide. In addition, the effect of 5 M guanidine-HCl on the conformation of fragments D1 and DD, which is known to abolish their antipolymerizing activity, was studied. Radioiodinated fibrinogen was used in the determination of Kconf, and quantitative analytical parameters, CI50% and CIs, calculated from competition between 125I-fibrinogen and the fibrinogen derivatives under study for binding to the immunochemically purified antibody. The measurements indicated that the epitope is unperturbed by iodination of fibrinogen and that 38.3% of fragment D1, 8.9% of fragment DD, 3.6% of the γ chain and less than 0.008% of the γ391-405 molecules adopt in aqueous solution the native conformation within the epitope. Denaturation of fragment D1 with 5 M guanidine-HCl affected only slightly the conformation of this γ chain determinant. More significant changes in the conformation were observed when fragment DD was denatured. The results 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 which is in close vicinity to the polymerization site, crosslinking site, and platelet recognition site.

Effect of Calcium and Synthetic Peptides on Fibrin Polymerization

1982 ◽  
Vol 47 (02) ◽  
pp. 118-121 ◽  
Author(s):  
M Furlan ◽  
C Rupp ◽  
E A Beck ◽  
L Svendsen
Keyword(s):  
Conformational Changes ◽  
Binding Sites ◽  
Synthetic Peptide ◽  
Calcium Concentration ◽  
Polymerization Process ◽  
Amino Terminus ◽  
Human Fibrinogen ◽  
Agkistrodon Contortrix ◽  
Β Chain ◽  
Fibrin Monomers

SummaryHuman fibrinogen was subjected to limited proteolytic attack by thrombin, batroxobin or Agkistrodon contortrix thrombin-like enzyme, yielding desAB-, desA- or desB-fibrin monomers, respectively. Turbidity curves demonstrated that, with all three enzymes, the polymerization process was strongly accelerated by increasing the calcium concentration from 10−5 M to 10−4 M. Synthetic peptide Gly-His-Arg (5 mM), an analogue of the aminoterminal sequence of fibrin β-chain, inhibited aggregation of desB-fibrin monomers at physiological calcium concentration whereas it enhanced aggregation of desA- and desAB-fibrin monomers at calcium concentrations below 10−4 M. On the other hand, Gly-Pro-Arg (1 mM) corresponding to the amino-terminus of fibrin α-chain, dramatically inhibited aggregation of both desA- and desB-fibrins, but it only moderately affected the polymerization of thrombin-induced monomers. We conclude that the observed effects of Gly-Pro-Arg and Gly-His-Arg are not due solely to their competition with fibrin amino-termini for the respective binding sites in the D-domain, but rather reflect conformational changes in fibrin monomers which affect the polymerization process.

Conformation of Ribosomes from Escherichia Coli

1974 ◽  
Vol 32 ◽  
pp. 210-211
Author(s):  
M. Boublik ◽  
W. Hellmann ◽  
F. Jenkins
Keyword(s):  
Binding Sites ◽  
Ribosomal Proteins ◽  
Fluorescent Dyes ◽  
Protein Biosynthesis ◽  
Three Dimensional ◽  
Spatial Arrangement ◽  
Dimensional Structure ◽  
Complete Understanding ◽  
And Function

The present knowledge of the three-dimensional structure of ribosomes is far too limited to enable a complete understanding of the various roles which ribosomes play in protein biosynthesis. The spatial arrangement of proteins and ribonuclec acids in ribosomes can be analysed in many ways. Determination of binding sites for individual proteins on ribonuclec acid and locations of the mutual positions of proteins on the ribosome using labeling with fluorescent dyes, cross-linking reagents, neutron-diffraction or antibodies against ribosomal proteins seem to be most successful approaches. Structure and function of ribosomes can be correlated be depleting the complete ribosomes of some proteins to the functionally inactive core and by subsequent partial reconstitution in order to regain active ribosomal particles.

Determination of copper binding sites in peptides containing basic residues: a combined experimental and theoretical study

2001 ◽  
Vol 204 (1-3) ◽  
pp. 31-46 ◽  
Author(s):  
Brian K. Bluhm ◽  
Sharon J. Shields ◽  
Craig A. Bayse ◽  
Michael B. Hall ◽  
David H. Russell
Keyword(s):  
Binding Sites ◽  
Theoretical Study ◽  
Copper Binding ◽  
Determination Of Copper ◽  
Copper Binding Sites

scholarly journals Determination of the alpha-actinin-binding site on actin filaments by cryoelectron microscopy and image analysis.

1994 ◽  
Vol 126 (2) ◽  
pp. 433-443 ◽  
Author(s):  
A McGough ◽  
M Way ◽  
D DeRosier
Keyword(s):  
Image Analysis ◽  
Smooth Muscle ◽  
Binding Sites ◽  
Actin Filaments ◽  
Three Dimensional ◽  
Actin Binding ◽  
Dimensional Structure ◽  
Outer Face ◽  
Actin Binding Domain

The three-dimensional structure of actin filaments decorated with the actin-binding domain of chick smooth muscle alpha-actinin (alpha A1-2) has been determined to 21-A resolution. The shape and location of alpha A1-2 was determined by subtracting maps of F-actin from the reconstruction of decorated filaments. alpha A1-2 resembles a bell that measures approximately 38 A at its base and extends 42 A from its base to its tip. In decorated filaments, the base of alpha A1-2 is centered about the outer face of subdomain 2 of actin and contacts subdomain 1 of two neighboring monomers along the long-pitch (two-start) helical strands. Using the atomic model of F-actin (Lorenz, M., D. Popp, and K. C. Holmes. 1993. J. Mol. Biol. 234:826-836.), we have been able to test directly the likelihood that specific actin residues, which have been previously identified by others, interact with alpha A1-2. Our results indicate that residues 86-117 and 350-375 comprise distinct binding sites for alpha-actinin on adjacent actin monomers.

Determination of spatial domains of zygotic gene expression in the Drosophila embryo by the affinity of binding sites for the bicoid morphogen

Nature ◽  
1989 ◽  
Vol 340 (6232) ◽  
pp. 363-367 ◽  
Author(s):  
Wolfgang Driever ◽  
Gudrun Thoma ◽  
Christiane Nüsslein-Volhard
Keyword(s):  
Gene Expression ◽  
Binding Sites ◽  
Drosophila Embryo ◽  
Zygotic Gene ◽  
Spatial Domains

Modelling competition between the siderophore ferrioxamine B and humic like binding sites in seawater v1

2021 ◽  
Author(s):  
Kechen Zhu ◽  
Martha Gledhill
Keyword(s):  
Binding Sites ◽  
Major Ions ◽  
Side Reaction ◽  
Iron Solubility ◽  
Desferrioxamine B ◽  
Thermodynamic Constants ◽  
Reaction Coefficient ◽  
Ferrioxamine B ◽  
Donnan Model

Here we archive a protocol that can be used to determine competition between a siderophore (ferrioxamine B) and humic like binding sites that are present in marine DOM. We use the NICA-Donnan model to describe binding by humic like binding sites in DOM. Constants for Fe binding to marine DOM are taken from Zhu et al., (2021). Thermodynamic constants describing binding between major ions, iron and ferrioxamine B are taken from Schijf and Burns, (2016). References Schijf, J., Burns, S.M., 2016. Determination of the Side-Reaction Coefficient of Desferrioxamine B in Trace-Metal-Free Seawater. Front. Mar. Sci. 3, 117. https://doi.org/10.3389/fmars.2016.00117 Zhu, K., Birchill, A.J., Milne, A., Ussher, S.J., Humphreys, M.P., Carr, N., Mahaffey, C., Lohan, M.C., Achterberg, E.P., Gledhill, M., 2021a. Equilbrium calculations of iron speciation and apparent iron solubility in the Celtic Sea at ambient pH using the NICA-Donnan model. Mar. Chem

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