Enhancement of Plasminogen Binding to U937 Cells and Fibrin by Complestatin

1997 ◽  
Vol 77 (01) ◽  
pp. 137-142 ◽  
Author(s):  
Kiyoshi Tachikawa ◽  
Keiji Hasurni ◽  
Akira Endo
Keyword(s):  
Binding Site ◽  
Binding Affinity ◽  
Blood Cells ◽  
Aminocaproic Acid ◽  
U937 Cells ◽  
Biochemical Basis ◽  
Equilibrium Binding ◽  
Pharmacological Stimulation ◽  
Endogenous Fibrinolysis ◽  
Stimulation Of

SummaryPlasminogen binds to endothelial and blood cells as well as to fibrin, where the zymogen is efficiently activated and protected from inhibition by α2-antiplasmin. In the present study we have found that complestatin, a peptide-like metabolite of a streptomyces, enhances binding of plasminogen to cells and fibrin. Complestatin, at concentrations ranging from 1 to 5 μM, doubled 125I-plasminogen binding to U937 cells both in the absence and presence of lipoprotein(a), a putative physiological competitor of plasminogen. The binding of 125I-plasminogen in the presence of complestatin was abolished by e-aminocaproic acid, suggesting that the lysine binding site(s) of the plasminogen molecule are involved in the binding. Equilibrium binding analyses indicated that complestatin increased the maximum binding of 125I-plasminogen to U937 cells without affecting the binding affinity. Complestatin was also effective in increasing 125I-plasminogen binding to fibrin, causing 2-fold elevation of the binding at ~1 μM. Along with the potentiation of plasminogen binding, complestatin enhanced plasmin formation, and thereby increased fibrinolysis. These results would provide a biochemical basis for a pharmacological stimulation of endogenous fibrinolysis through a promotion of plasminogen binding to cells and fibrin.

SCFβ-TRCP Mediates Ubiquitination and Degradation of the Erythropoietin Receptor and Controls Cell Proliferation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 464-464
Author(s):  
Frederique Verdier ◽  
Laure Meyer ◽  
Benedicte Deau ◽  
Hana Forejtnikova ◽  
Dominique Dumenil ◽  
...  
Keyword(s):  
Binding Site ◽  
Ubiquitin Ligase ◽  
Blood Cells ◽  
E3 Ubiquitin Ligase ◽  
Ubiquitin Proteasome System ◽  
Erythropoietin Receptor ◽  
Erythroid Progenitors ◽  
Ubiquitin Proteasome ◽  
Binding Sequence ◽  
Stimulation Of

Abstract Control of intensity and duration of erythropoietin (Epo) signalling is necessary to tightly regulate red blood cells production. After Epo stimulation of erythroid cells, 2 types of signal are transduced via the Epo receptor (Epo-R): positive signals involved in survival and proliferation, and negative signals involved in signal arrest. We have recently shown that the ubiquitin/ proteasome system plays a major role in the control of Epo-R signalling duration and desensitisation processes. Indeed, after Epo stimulation the Epo-R is ubiquitinated and its intracellular part is degraded by the proteasome, preventing further signal transduction. The remaining part of the receptor, together with associated Epo is internalised and degraded by the lysosomes (Walrafen et al 2005 Blood, 105, 600-608). Our aim was to identify the E3 ubiquitin ligase involved in Epo-R ubiquitination. The Epo-R contains a putative β-Trcp binding site in its intracellular domain. Interestingly, this putative binding sequence is located in a region of the Epo-R that is deleted in erythroid progenitors from patients with familial polycythemia. We show that β-Trcp is responsible for Epo-R ubiquitination upon Epo stimulation. After Epo stimulation, β-Trcp binds to the Epo-R and this binding is dependent on Jak2 activation. Mutation of the Ser 462 residue of the Epo-R, located in the consensus β-Trcp binding site abolished β-Trcp binding, Epo-R ubiquitination and EpoR cleavage by the proteasome. Activation of the mutated Epo-R is prolonged in comparaison with Epo-R WT and BaF3 cells expressing this mutated receptor unable to bind β-Trcp are hypersensitive to Epo. Whether the removal of the β-Trcp binding site contributes to the hypersensitivity to Epo in familial polycythemia is currently under study.]

scholarly journals Excision and transposition of Tn5 as an SOS activity in Escherichia coli.

Genetics ◽  
1991 ◽  
Vol 128 (1) ◽  
pp. 45-57 ◽  
Author(s):  
C T Kuan ◽  
S K Liu ◽  
I Tessman
Keyword(s):  
Escherichia Coli ◽  
Binding Site ◽  
Reca Protein ◽  
Precursor Molecule ◽  
Functional Integrity ◽  
Lexa Protein ◽  
Lexa Binding ◽  
Additional Role ◽  
Stimulation Of

Abstract Excision and transposition of the Tn5 element in Escherichia coli ordinarily appear to occur by recA-independent mechanisms. However, recA(Prtc) genes, which encode RecA proteins that are constitutively activated to the protease state, greatly enhanced excision and transposition; both events appeared to occur concomitantly and without destruction of the donor DNA. The recombinase function of the RecA protein was not required. Transposition was accompanied by partial, and occasionally full, restoration of the functional integrity of the gene vacated by the excised Tn5. The stimulation of transposition was inhibited by an uncleavable LexA protein and was strongly enhanced by an additional role of the RecA(Prtc) protein besides its mediation of LexA cleavage. To account for the enhanced transposition, we suggest that (i) there may be a LexA binding site within the promoter for the IS50 transposase, (ii) activated RecA may cleave the IS50 transposition inhibitor, and (iii) the transposase may be formed by RecA cleavage of a precursor molecule.

scholarly journals Identification of Cytokines in Whole Blood for Differential Diagnosis of Tuberculosis versus Pneumonia

2010 ◽  
Vol 17 (5) ◽  
pp. 771-777 ◽  
Author(s):  
Wen-Lin Su ◽  
Wann-Cherng Perng ◽  
Ching-Hui Huang ◽  
Cheng-Yu Yang ◽  
Chin-Pyng Wu ◽  
...  
Keyword(s):  
Whole Blood ◽  
Blood Cells ◽  
Cutoff Point ◽  
Interleukin 12 ◽  
Operating Characteristics ◽  
Percent Change ◽  
Ifn Γ ◽  
Sensitivity Specificity ◽  
Whole Blood Cells ◽  
Stimulation Of

ABSTRACT Differentiating tuberculosis (TB) from pneumonia remains a challenge. We evaluated the cytokine profiles of whole blood cells from patients with TB (n = 38) or pneumonia (n = 30) and from healthy individuals (n = 30) before and after stimulating cells with ESAT-6 or lipopolysaccharide (LPS). When the percent change in the levels of gamma interferon (IFN-γ) after stimulation with ESAT-6 was used in receiver operating characteristics (ROC) analysis (a graphic method to determine the diagnostic accuracy of a test) to identify a patient with TB, the area under the curve (AUC) was 90.4%, and a cutoff point of a 3.59% change produced a corresponding sensitivity, specificity, and accuracy of over 80%. When the change in IFN-γ after stimulation of blood cells with LPS was used to identify a patient with pneumonia, the AUC reached 89.1%, and a cutoff point of 3.59% produced a sensitivity, specificity, and accuracy of approximately 80% each. When the change in interleukin-12 (IL-12) after stimulation of blood cells with LPS was selected to define a patient with pneumonia, the AUC was 85.2%, and a cutoff point of 2.08% gave a sensitivity, specificity, and accuracy of 80.0%, 78.9%, and 79.4%, respectively. We conclude that the percent change in IFN-γ after stimulation of whole blood cells with ESAT-6 may differentiate patients with TB from patients with pneumonia. The percent change in IFN-γ and IL-12 after LPS stimulation of whole blood cells could differentiate patients with pneumonia from patients with TB.

scholarly journals Effects of Selected Prebiotics or Synbiotics Administered in ovo on Lymphocyte Subsets in Bursa of the Fabricius, Thymus, and Spleen in Non-Immunized and Immunized Chicken Broilers

Animals ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 476
Author(s):  
Marianna Szczypka ◽  
Agnieszka Suszko-Pawłowska ◽  
Maciej Kuczkowski ◽  
Michał Gorczykowski ◽  
Magdalena Lis ◽  
...  
Keyword(s):  
Immune System ◽  
Lactococcus Lactis ◽  
Blood Cells ◽  
Broiler Chickens ◽  
Lymphocyte Subsets ◽  
Double Positive ◽  
In Ovo ◽  
Lactococcus Lactis Subsp ◽  
Egg Incubation ◽  
Stimulation Of

The effects of in ovo-delivered prebiotics and synbiotics on the lymphocyte subsets of the lymphoid organs in non-immunized 7-day-old broiler chickens and in non-immunized, sheep red blood cells (SRBC)-immunized, and dextran (DEX)-immunized 21- and 35-day-old birds were studied. The substances were injected on the 12th day of egg incubation: Prebiotic1 group (Pre1) with a solution of inulin, Prebiotic2 group (Pre2) with a solution of Bi2tos (non-digestive transgalacto-oligosaccharides), Synbiotic1 group (Syn1) with inulin and Lactococcus lactis subsp. lactis IBB SL1, and Synbiotic2 group (Syn2) with Bi2tos and Lactococcus lactis subsp. cremoris IBB SC1. In 7-day-old chicks, a decrease in T splenocytes was noticed in all groups. The most pronounced effect in 21- and 35-day-old birds was an increase in TCRγδ+ cells in Syn1 and Syn2 groups. A decrease in bursal B cells was observed in DEX-immunized Pre1 group (21-day-old birds), and in the Syn1 group in non-immunized and SRBC-immunized 35-day-old birds. An increase in double-positive lymphocytes was observed in Pre1 (35-day-old birds) and Pre2 (immunized 21-day-old birds) groups. In Pre1 and Syn1 groups (21- and 35-day-old), an increase in B splenocytes and a decrease in T splenocytes were observed. We concluded that Syn1 was the most effective in the stimulation of the chicken immune system.

C-fos expression in the rat brain after pharmacological stimulation of the rat “mediodorsal” thalamus by means of microdialysis

Neuroscience ◽  
1995 ◽  
Vol 66 (1) ◽  
pp. 115-131 ◽  
Author(s):  
E.B.H.W. Erdtsieck-Ernste ◽  
M.G.P. Feenstra ◽  
M.H.A. Botterblom ◽  
H.F.M. Van Uum ◽  
A.A. Sluiter ◽  
...  
Keyword(s):  
Rat Brain ◽  
Mediodorsal Thalamus ◽  
Pharmacological Stimulation ◽  
Fos Expression ◽  
Stimulation Of

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 High resolution structure of human apolipoprotein (a) kringle IV type 2: beyond the lysine binding site

2020 ◽  
Vol 61 (12) ◽  
pp. 1687-1696
Author(s):  
Alice Santonastaso ◽  
Maristella Maggi ◽  
Hugo De Jonge ◽  
Claudia Scotti
Keyword(s):  
High Resolution ◽  
Binding Site ◽  
Small Molecule ◽  
Aminocaproic Acid ◽  
Variable Number ◽  
Crystallographic Structure ◽  
Heparin Binding ◽  
Human Apolipoprotein ◽  
Unique Shape

Lipoprotein (a) [Lp(a)] is characterized by an LDL-like composition in terms of lipids and apoB100, and by one copy of a unique glycoprotein, apo(a). The apo(a) structure is mainly based on the repetition of tandem kringle domains with high homology to plasminogen kringles 4 and 5. Among them, kringle IV type 2 (KIV-2) is present in a highly variable number of genetically encoded repeats, whose length is inversely related to Lp(a) plasma concentration and cardiovascular risk. Despite it being the major component of apo(a), the actual function of KIV-2 is still unclear. Here, we describe the first high-resolution crystallographic structure of this domain. It shows a general fold very similar to other KIV domains with high and intermediate affinity for the lysine analog, ε-aminocaproic acid. Interestingly, KIV-2 presents a lysine binding site (LBS) with a unique shape and charge distribution. KIV-2 affinity for predicted small molecule binders was found to be negligible in surface plasmon resonance experiments; and with the LBS being nonfunctional, we propose to rename it “pseudo-LBS”. Further investigation of the protein by computational small-molecule docking allowed us to identify a possible heparin-binding site away from the LBS, which was confirmed by specific reverse charge mutations abolishing heparin binding. This study opens new possibilities to define the pathogenesis of Lp(a)-related diseases and to facilitate the design of specific therapeutic drugs.

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