scholarly journals A synthetic peptide derived from the carboxy terminus of the laminin A chain represents a binding site for the alpha 3 beta 1 integrin

1992 ◽  
Vol 117 (2) ◽  
pp. 449-459 ◽  
Author(s):  
KR Gehlsen ◽  
P Sriramarao ◽  
LT Furcht ◽  
AP Skubitz
Keyword(s):  
Cell Adhesion ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Binding Site ◽  
Synthetic Peptide ◽  
Beta 1 Integrin ◽  
Carboxy Terminal Region ◽  
A Chain ◽  
Coated Surfaces ◽  
Carboxy Terminal

The purpose of this study was to identify the binding site(s) within laminin for the alpha 3 beta 1 integrin receptor. It has been previously shown, using proteolytic fragments and anti-laminin antibodies, that the region in laminin for alpha 3 beta 1 integrin binding is localized to the carboxy-terminal region at the end of the long arm (Gehlsen, K. R., E. Engvall, K. Dickerson, W. S. Argraves, and E. Ruoslahti. 1989. J. Biol. Chem. 264:19034-19038; Tomaselli, K. J., D. E. Hall, L. T. Reichardt, L. A. Flier, K. R. Gehlsen, D. C. Turner, and S. Carbonetto. 1990. Neuron. 5:651-662). Using synthetic peptides, we have identified an amino acid sequence within the carboxy-terminal region of the laminin A chain that is recognized by the alpha 3 beta 1 integrin. The amino acid sequence represented by the synthetic peptide GD-6 (KQNCLSSRASFRGCVRNLRLSR residues numbered 3011 to 3032) of the globular domain of the murine A chain supports cell attachment and inhibits cell adhesion to laminin-coated surfaces. By affinity chromatography, peptide GD-6-Sepharose specifically bound solubilized alpha 3 beta 1 from extracts of surface-iodinated cells in a cation-dependent manner, while it did not bind other integrins. In addition, exogenous peptide GD-6 specifically eluted bound alpha 3 beta 1 from laminin-Sepharose columns but did not elute the alpha 3 beta 1 integrin from a fibronectin-Sepharose column. Using integrin subunit-specific monoclonal antibodies, only those antibodies against the alpha 3 and beta 1 subunits inhibited cell adhesion to peptide GD-6-coated surfaces. Finally, a polyclonal antibody made against peptide GD-6 reacted specifically with both murine and human laminin and significantly inhibited cell adhesion to laminin-coated surfaces but not those coated with other matrix proteins. These results identify the laminin A chain amino acid sequence of peptide GD-6 as representing a binding site in laminin for the alpha 3 beta 1 integrin.

The Amino Acid Sequence in Fibrin Responsible for High Affinity Thrombin Binding

2001 ◽  
Vol 85 (03) ◽  
pp. 470-474 ◽  
Author(s):  
Kevin Siebenlist ◽  
Stephen Brennan ◽  
Trudy Holyst ◽  
Michael Mosesson ◽  
David Meh
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Binding Site ◽  
Binding Affinity ◽  
Ionization Mass ◽  
Consensus Sequences ◽  
High Affinity ◽  
High Affinity Binding Site ◽  
Carboxy Terminal ◽  
Sulfated Peptides

SummaryHuman fibrin has a low affinity thrombin binding site in its E domain and a high affinity binding site in the carboxy-terminal region of its variant ’ chain (’408-427). Comparison of the ’ amino acid sequence (VRPEHPAETEYDSLYPEDDL) with other protein sequences known to bind to thrombin exosites such as those in GPIb , the platelet thrombin receptor, thrombomodulin, and hirudin suggests no homology or consensus sequences, but Glu and Asp enrichment are common to all. Tyrosine sulfation in these sequences enhances thrombin exosite binding, but this has not been uniformly investigated. The fibrinogen ’ chain mass determined by electrospray ionization mass spectrometry, was 50,549 Da, a value 151 Da greater than predicted from its amino acid/carbohydrate sequence. Since each sulfate group increases mass by 80 Da, this indicates that both tyrosines at 418 and 422 are sulfated. A series of overlapping ’ peptides was prepared for evaluation of their inhibition of 125I-labeled PPACK-thrombin binding to fibrin. ’414-427 was as effective an inhibitor as ’408-427 and its binding affinity was dependent on all carboxy-terminal residues. Mono Tyr-sulfated peptides were prepared by substituting non-sulfatable Phe for Tyr at ’ 418 or 422. Sulfation at either Tyr residue increased binding competition compared with non-sulfated peptides, but was less effective than doubly sulfated peptides, which had 4 to 8-fold greater affinity. The reverse ’ peptide or the forward sequence with repositioned Tyr residues did not compete well for thrombin binding, indicating that the positions of charged residues are important for thrombin binding affinity

scholarly journals Ankyrin-binding proteins related to nervous system cell adhesion molecules: candidates to provide transmembrane and intercellular connections in adult brain.

1993 ◽  
Vol 121 (1) ◽  
pp. 121-133 ◽  
Author(s):  
J Q Davis ◽  
T McLaughlin ◽  
V Bennett
Keyword(s):  
Nervous System ◽  
Cell Adhesion ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Binding Site ◽  
Adhesion Molecules ◽  
Cell Adhesion Molecules ◽  
Binding Proteins ◽  
Cytoplasmic Domain ◽  
System Cell

A major class of ankyrin-binding glycoproteins have been identified in adult rat brain of 186, 155, and 140 kD that are alternatively spliced products of the same pre-mRNA. Characterization of cDNAs demonstrated that ankyrin-binding glycoproteins (ABGPs) share 72% amino acid sequence identity with chicken neurofascin, a membrane-spanning neural cell adhesion molecule in the Ig super-family expressed in embryonic brain. ABGP polypeptides have the following features consistent with a role as ankyrin-binding proteins in vitro and in vivo: (a) ABGPs and ankyrin associate as pure proteins in a 1:1 molar stoichiometry; (b) the ankyrin-binding site is located in the COOH-terminal 21 kD of ABGP186 which contains the predicted cytoplasmic domain; (c) ABGP186 is expressed at approximately the same levels as ankyrin (15 pmoles/milligram of membrane protein); and (d) ABGP polypeptides are co-expressed with the adult form of ankyrinB late in postnatal development and are colocalized with ankyrinB by immunofluorescence. Similarity in amino acid sequence and conservation of sites of alternative splicing indicate that genes encoding ABGPs and neurofascin share a common ancestor. However, the major differences in developmental expression reported for neurofascin in embryos versus the late postnatal expression of ABGPs suggest that ABGPs and neurofascin represent products of gene duplication events that have subsequently evolved in parallel with distinct roles. The predicted cytoplasmic domains of rat ABGPs and chicken neurofascin are nearly identical to each other and closely related to a group of nervous system cell adhesion molecules with variable extracellular domains, which includes L1, Nr-CAM, and Ng-CAM of vertebrates, and neuroglian of Drosophila. The ankyrin-binding site of rat ABGPs is localized to the C-terminal 200 residues which encompass the cytoplasmic domain, suggesting the hypothesis that ability to associate with ankyrin may be a shared feature of neurofascin and related nervous system cell adhesion molecules.

scholarly journals Synthetic peptides from the carboxy-terminal globular domain of the A chain of laminin: their ability to promote cell adhesion and neurite outgrowth, and interact with heparin and the beta 1 integrin subunit.

1991 ◽  
Vol 115 (4) ◽  
pp. 1137-1148 ◽  
Author(s):  
A P Skubitz ◽  
P C Letourneau ◽  
E Wayner ◽  
L T Furcht
Keyword(s):  
Amino Acids ◽  
Cell Adhesion ◽  
Neurite Outgrowth ◽  
Cell Types ◽  
Integrin Subunit ◽  
Globular Domain ◽  
Heparin Binding ◽  
Beta 1 Integrin ◽  
A Chain ◽  
Carboxy Terminal

The large carboxy-terminal globular domain (G domain; residues 2,110-3,060) of the A chain of murine-derived laminin has been shown to promote heparin binding, cell adhesion, and neurite outgrowth. This study was conducted to define the potential sequence(s) originating from the G domain of laminin with any of these functional activities. A series of peptides were synthesized from the G domain, termed GD peptides, each approximately 20 amino acids long and containing multiple positively charged amino acids. In direct 3H-heparin binding assays, peptides GD-1 and GD-2 bound high levels of 3H-heparin, while peptides GD-3 and GD-4 bound lower levels of 3H-heparin, and GD-5 bound essentially no 3H-heparin. The binding of 3H-heparin to peptides GD-1 and GD-2 appeared to be of high affinity, since significant binding of 3H-heparin to these two peptides was still observed even when the NaCl concentration was raised to 1.0 M. Four of the peptides, GD-1, GD-2, GD-3, and GD-4, directly promoted the adhesion and spreading of HT-1080 human fibrosarcoma cells as well as the outgrowth of neurites from chick spinal cord and dorsal root ganglia neurons. In addition, solutions of these peptides or antibodies generated against these peptides inhibited laminin-mediated HT-1080 cell adhesion. Antibodies against the beta 1 integrin subunit inhibited HT-1080 cell adhesion and neurite outgrowth on surfaces adsorbed with peptides GD-3 and GD-4. Therefore, laminin appears to have multiple, independent sequences in the G domain that serve a similar cell adhesion promoting function for different cell types. Furthermore, these results suggest that the sequences comprising peptides GD-3 and GD-4 use an integrin as a receptor, of which the beta 1 integrin subunit is a component for these various cell types.

scholarly journals Sequence of a highly divergent beta tubulin gene reveals regional heterogeneity in the beta tubulin polypeptide.

1984 ◽  
Vol 99 (5) ◽  
pp. 1754-1760 ◽  
Author(s):  
K F Sullivan ◽  
D W Cleveland
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Predicted Amino Acid Sequence ◽  
Tubulin Gene ◽  
Beta Tubulin ◽  
Protein Coding ◽  
Evolutionary Selection ◽  
Carboxy Terminal Region ◽  
Beta 2 ◽  
Carboxy Terminal

The nucleotide sequence of a chicken genomic DNA segment containing the chicken beta 4 tubulin gene has been determined. The predicted amino acid sequence of beta 4 is surprisingly divergent from that of the chicken beta 2 gene that encodes the dominant neural beta tubulin. beta 4 differs from beta 2 at 36 residue positions and encodes a polypeptide that is four amino acids longer, yielding a divergence of 8.9% between the two beta tubulin isotypes. While many of the amino acid substitutions are conservative, several involve significant alteration in the physiochemical properties of the residue. Furthermore, the amino acid substitution positions are not randomly located within the primary sequence but are distinctly clustered: major divergence occurs in the carboxy-terminal region beyond residue 430 and within the second protein coding exon segments of the genes. In addition, large regions of absolute sequence conservation are also present. Certain sequences within the heterogeneous regions are conserved in other species, indicating that these regions are under positive evolutionary selection pressure and are therefore probably essential for some aspect of beta-tubulin function. These findings strongly suggest that regional amino acid sequence heterogeneity may play an important role in the establishment of functionally differentiated beta tubulin polypeptides.

Amino acid sequence of the A-chain of the mistletoe lectin I

Peptides 1994 ◽  
1995 ◽  
pp. 737-738
Author(s):  
M. Huguet ◽  
S. Stoeva ◽  
C. Decker ◽  
S. Wilhelm ◽  
T. Stiefel ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Mistletoe Lectin ◽  
A Chain

Plasmin-platelet interaction involves cleavage of functional thrombin receptor

1996 ◽  
Vol 271 (1) ◽  
pp. C54-C60 ◽  
Author(s):  
M. Kimura ◽  
T. T. Andersen ◽  
J. W. Fenton ◽  
W. F. Bahou ◽  
A. Aviv
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Platelet Activation ◽  
Synthetic Peptide ◽  
Time Dependent ◽  
Thrombin Receptor ◽  
Amino Acid Residues ◽  
Enzymatic Action ◽  
High Concentrations ◽  
Inhibition Of Thrombin

We tested the hypothesis that the inhibition of thrombin-induced platelet activation by plasmin is mediated via the enzymatic action of plasmin on the functional thrombin receptor. We monitored the binding of the anti-thrombin receptor antibody [anti-TR-(34-46)] to platelets; this binding is sensitive to the cleavage of the thrombin receptor at amino acid residues Arg-41 to Ser-42. Plasmin inhibited anti-TR-(34-46) binding in dose- and time-dependent manners. The inactive synthetic peptide with the amino acid sequence 40-55 of the thrombin receptor (D-FPRSFLLRNPNDKYEPF) was similarly cleaved by thrombin and plasmin to an active peptide (SFLLRNPNDKYEPF) that produced robust cytosolic Ca2+ responses. At high concentrations, plasmin itself can activate platelets. We explored this effect with the use of anti-TR-(1-160). This antibody abolished the cytosolic Ca2+ responses to thrombin and to the thrombin receptor-activating peptide SFLLRN but did not attenuate the plasmin-induced cytosolic Ca2+ response. Thus plasmin inhibits thrombin-evoked platelet activation by cleaving the thrombin receptor, but the plasmin-induced cytosolic Ca2+ response is not due to the generation of the tethered peptide of the thrombin receptor.

Amino acid sequence of human thrombin A chain

1974 ◽  
Vol 60 (2) ◽  
pp. 717-722 ◽  
Author(s):  
Daniel A. Walz ◽  
Walter H. Seegers
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Human Thrombin ◽  
A Chain
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