scholarly journals Comparison of Target Recognition by TRAF1 and TRAF2

2020 ◽  
Vol 21 (8) ◽  
pp. 2895
Author(s):  
Chang Min Kim ◽  
Hyun Ho Park
Keyword(s):  
Receptor Binding ◽  
Target Recognition ◽  
Affinity Binding ◽  
Binding Affinities ◽  
Amino Acid Residues ◽  
Critical Determinant ◽  
High Affinity Binding ◽  
High Affinity ◽  
Caspase 2

Although TRAF1 and TRAF2 share common receptors and have extremely conserved amino acid residues, recent studies have shown that key differences in receptor binding preferences with different affinities exist, which might be important for their different functions in TRAF-mediated signal transduction. To better understand TRAF1 and TRAF2 signaling, we analyzed and compared their receptor binding-affinities. Our study revealed that TRADD, TANK, and caspase-2 bind to both TRAF1 and TRAF2 with different affinities in vitro. Sequence and structural analyses revealed that S454 on TRAF2 (corresponding to A369 of TRAF1) is critical for the binding of TRADD, and F347 on TRAF1 (corresponding to L432 of TRAF2) is a critical determinant for high affinity binding of TANK and caspase-2.

Shark Attack: High affinity binding proteins derived from shark vNAR domains by stepwise in vitro affinity maturation

2014 ◽  
Vol 191 ◽  
pp. 236-245 ◽  
Author(s):  
Stefan Zielonka ◽  
Niklas Weber ◽  
Stefan Becker ◽  
Achim Doerner ◽  
Andreas Christmann ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Affinity Maturation ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity

scholarly journals A broadly neutralizing antibody protects against SARS-CoV, pre-emergent bat CoVs, and SARS-CoV-2 variants in mice

2021 ◽  
Author(s):  
David R. Martinez ◽  
Alexandra Schaefer ◽  
Sophie Gobeil ◽  
Dapeng Li ◽  
Gabriela De la Cruz ◽  
...  
Keyword(s):  
Structural Analysis ◽  
Receptor Binding ◽  
Specific Antibody ◽  
Neutralizing Antibody ◽  
Affinity Binding ◽  
Receptor Binding Domain ◽  
High Affinity Binding ◽  
High Affinity ◽  
Broadly Neutralizing Antibody ◽  
Conserved Epitope

AbstractSARS-CoV in 2003, SARS-CoV-2 in 2019, and SARS-CoV-2 variants of concern (VOC) can cause deadly infections, underlining the importance of developing broadly effective countermeasures against Group 2B Sarbecoviruses, which could be key in the rapid prevention and mitigation of future zoonotic events. Here, we demonstrate the neutralization of SARS-CoV, bat CoVs WIV-1 and RsSHC014, and SARS-CoV-2 variants D614G, B.1.1.7, B.1.429, B1.351 by a receptor-binding domain (RBD)-specific antibody DH1047. Prophylactic and therapeutic treatment with DH1047 demonstrated protection against SARS-CoV, WIV-1, RsSHC014, and SARS-CoV-2 B1.351infection in mice. Binding and structural analysis showed high affinity binding of DH1047 to an epitope that is highly conserved among Sarbecoviruses. We conclude that DH1047 is a broadly neutralizing and protective antibody that can prevent infection and mitigate outbreaks caused by SARS-like strains and SARS-CoV-2 variants. Our results argue that the RBD conserved epitope bound by DH1047 is a rational target for pan Group 2B coronavirus vaccines.

scholarly journals Four Amino Acid Residues Are Critical for High Affinity Binding of Neuromedin B to the Neuromedin B Receptor

1998 ◽  
Vol 273 (26) ◽  
pp. 15927-15932 ◽  
Author(s):  
Eduardo Sainz ◽  
Mark Akeson ◽  
Samuel A. Mantey ◽  
Robert T. Jensen ◽  
James F. Battey
Keyword(s):  
Amino Acid ◽  
Affinity Binding ◽  
Amino Acid Residues ◽  
High Affinity Binding ◽  
High Affinity ◽  
Neuromedin B

scholarly journals Modification of the 85-kilodalton subunit of phosphatidylinositol-3 kinase in platelet-derived growth factor-stimulated cells.

1992 ◽  
Vol 12 (8) ◽  
pp. 3415-3424 ◽  
Author(s):  
W M Kavanaugh ◽  
A Klippel ◽  
J A Escobedo ◽  
L T Williams
Keyword(s):  
Tyrosine Phosphorylation ◽  
Platelet Derived Growth Factor ◽  
Affinity Binding ◽  
Pdgf Receptor ◽  
Phosphatidylinositol 3 Kinase ◽  
High Affinity Binding ◽  
High Affinity ◽  
Pdgf Stimulation ◽  
Stimulation Of

The activated platelet-derived growth factor (PDGF) receptor physically associates with p85, a subunit of phosphatidylinositol-3 kinase. Although this interaction may activate phosphatidylinositol-kinase and is crucial for PDGF-induced mitogenesis, it has not been shown whether p85 is modified in the process. p85 contains two SH2 (Src homology) domains, designated SH2-N and SH2-C. Recent experiments have shown that the SH2-C domain alone determines high-affinity binding of p85 to the PDGF receptor. The function of SH2-N, which binds receptors with lower affinity, is unknown. In this study, using a receptor-blotting technique, we find that p85 is modified by PDGF stimulation of intact cells. This modification involves inhibition of binding of the SH2-N region of p85 to the PDGF receptor. Studies with vanadate suggest that tyrosine phosphorylation of p85 is responsible for the modification of p85 detected by receptor blotting. Furthermore, recombinant p85 is modified in a similar manner when it is tyrosine phosphorylated in vitro by PDGF receptors. Tyrosine phosphorylation of p85 does not block binding of the SH2-C domain and therefore does not release p85 from high-affinity binding sites on the receptor in vitro. Instead, phosphorylation may regulate the ability of the SH2-N of p85 to bind to a different portion of the PDGF receptor or to another molecule in the signaling complex. This study provides the first evidence that p85 is tyrosine phosphorylated upon PDGF stimulation of cells and suggests that tyrosine phosphorylation of p85 regulates its activity or its interaction with other proteins.

scholarly journals CDK9 Autophosphorylation Regulates High-Affinity Binding of the Human Immunodeficiency Virus Type 1 Tat–P-TEFb Complex to TAR RNA

2000 ◽  
Vol 20 (18) ◽  
pp. 6958-6969 ◽  
Author(s):  
Mitchell E. Garber ◽  
Timothy P. Mayall ◽  
Eric M. Suess ◽  
Jill Meisenhelder ◽  
Nancy E. Thompson ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
Immunodeficiency Virus ◽  
Tar Rna ◽  
Cdk9 Phosphorylation ◽  
Hiv 1

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) Tat interacts with cyclin T1 (CycT1), a regulatory partner of CDK9 in the positive transcription elongation factor (P-TEFb) complex, and binds cooperatively with CycT1 to TAR RNA to recruit P-TEFb and promote transcription elongation. We show here that Tat also stimulates phosphorylation of affinity-purified core RNA polymerase II and glutathioneS-transferase–C-terminal-domain substrates by CycT1-CDK9, but not CycH-CDK7, in vitro. Interestingly, incubation of recombinant Tat–P-TEFb complexes with ATP enhanced binding to TAR RNA dramatically, and the C-terminal half of CycT1 masked binding of Tat to TAR RNA in the absence of ATP. ATP incubation lead to autophosphorylation of CDK9 at multiple C-terminal Ser and Thr residues, and full-length CycT1 (amino acids 728) [CycT1(1–728)], but not truncated CycT1(1–303), was also phosphorylated by CDK9. P-TEFb complexes containing a catalytically inactive CDK9 mutant (D167N) bound TAR RNA weakly and independently of ATP, as did a C-terminal truncated CDK9 mutant that was catalytically active but unable to undergo autophosphorylation. Analysis of different Tat proteins revealed that the 101-amino-acid SF2 HIV-1 Tat was unable to bind TAR with CycT1(1–303) in the absence of phosphorylated CDK9, whereas unphosphorylated CDK9 strongly blocked binding of HIV-2 Tat to TAR RNA in a manner that was reversed upon autophosphorylation. Replacement of CDK9 phosphorylation sites with negatively charged residues restored binding of CycT1(1–303)-D167N-Tat, and rendered D167N a more potent inhibitor of transcription in vitro. Taken together, these results demonstrate that CDK9 phosphorylation is required for high-affinity binding of Tat–P-TEFb to TAR RNA and that the state of P-TEFb phosphorylation may regulate Tat transactivation in vivo.

Specific high-affinity binding of 125I-labeled mouse interferon to interferon resistant embryonal carcinoma cells in vitro

Virology ◽  
1981 ◽  
Vol 114 (2) ◽  
pp. 585-588 ◽  
Author(s):  
Michel Aguet ◽  
Ion Gresser ◽  
Ara G. Hovanessian ◽  
Marie-Thérèse Bandu ◽  
Brigitte Blanchard ◽  
...  
Keyword(s):  
Embryonal Carcinoma ◽  
Carcinoma Cells ◽  
Affinity Binding ◽  
Embryonal Carcinoma Cells ◽  
High Affinity Binding ◽  
High Affinity

scholarly journals Mutations in the DG Loop of Adenovirus Type 5 Fiber Knob Protein Abolish High-Affinity Binding to Its Cellular Receptor CAR

1999 ◽  
Vol 73 (11) ◽  
pp. 9508-9514 ◽  
Author(s):  
Ian Kirby ◽  
Elizabeth Davison ◽  
Andrew J. Beavil ◽  
Cecilia P. C. Soh ◽  
Thomas J. Wickham ◽  
...  
Keyword(s):  
Secondary Structure ◽  
Receptor Binding ◽  
Adenovirus Type ◽  
Cellular Receptor ◽  
Affinity Binding ◽  
Cd Spectra ◽  
Binding Studies ◽  
High Affinity Binding ◽  
High Affinity ◽  
Adenovirus Type 5

ABSTRACT The amino acid residues in adenovirus type 5 (Ad5) fiber that interact with its cellular receptor, the coxsackie B virus and Ad receptor (CAR), have not been defined. To investigate this, multiple mutations were constructed in the region between residues 479 and 497 in Ad5 fiber (β-strands E and F and the adjacent region of the DG loop). The effects of these mutations on binding to CAR were determined by use of cell-binding competition experiments, surface plasmon resonance, and direct binding studies. The mutation effects on the overall folding and secondary structure of the protein were assessed by circular dichroism (CD) spectroscopy. Deletions of two consecutive amino acids between residues 485 and 493 abolished high-affinity binding to CAR; the CD spectra indicated that although there was no disruption of the overall folding and secondary structure of the protein, local conformational changes did occur. Moreover, single site mutations in this region of residues with exposed, surface-accessible side chains, such as Thr492, Asn493, and Val495, had no effect on receptor binding, which demonstrates that these residues are not in contact with CAR themselves. This implies the involvement of residues in neighboring loop regions. Replacement of the segment containing the two very short β-strands E and F and the turn between them (residues 479 to 486) with the corresponding sequence from Ad3 (βEFAd3→5 mutation) resulted in the loss of receptor binding. The identical CD spectra for βEFAd3→5 and wild-type proteins suggest that these substitutions caused no conformational rearrangement and that the loss of binding may thus be due to the substitution of one or more critical contact residues. These findings have implications for our understanding of the interaction of Ad5 fiber with CAR and for the construction of targeted recombinant Ad5 vectors for gene therapy purposes.

Zinc Trafficking 1. Probing the Roles of Proteome, Metallothionein, and Glutathione

Metallomics ◽  
2021 ◽  
Author(s):  
Afsana Mahim ◽  
Mohammad Mahim ◽  
David H Petering
Keyword(s):  
Carbonic Anhydrase ◽  
Binding Sites ◽  
Affinity Binding ◽  
Conditional Stability ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Conditional Stability Constant ◽  
High Affinity Binding ◽  
High Affinity

Abstract The cellular trafficking pathways that conduct zinc to its sites of binding in functional proteins remain largely unspecified. In this study, the hypothesis was investigated that non-specific proteomic binding sites serve as intermediates in zinc trafficking. Proteome from pig kidney LLC-PK1 cells contains a large concentration of such sites, displaying an average conditional stability constant of 1010-11, that are dependent on sulfhydryl ligands to achieve high affinity binding of zinc. As a result, the proteome competes effectively with induced metallothionein for Zn2+ upon exposure of cells to extracellular Zn2+ or during in vitro direct competition. The reaction of added Zn2+ bound to proteome with apo-carbonic anhydrase was examined as a potential model for intracellular zinc trafficking. The extent of this reaction was inversely dependent upon proteome concentration and under cellular conditions thought to be negligible. The rate of reaction was strictly first order in both Zn2+ and apo-carbonic anhydrase and also considered to be insignificant in cells. Adding the low molecular weight fraction of cell supernatant to the proteome markedly enhanced the speed of this reaction, a phenomenon dependent on the presence of glutathione. In agreement, inclusion of glutathione accelerated the reaction in a concentration-dependent manner. The implications of abundant high affinity binding sites for Zn2+ within the proteome are considered in relation to their interaction with glutathione in the efficient delivery of Zn2+ to functional binding sites and in the operation of fluorescent zinc sensors as a tool to observe zinc trafficking.

1-acetyl-7-deacetylforskolin: A potential non-specific inactive analog of forskolin for estimation of its specific high-affinity binding and adenylyl cyclase stimulation in vitro

Life Sciences ◽  
1995 ◽  
Vol 57 (14) ◽  
pp. 1367-1373 ◽  
Author(s):  
Tom Sasaki ◽  
Kohsuke Furukata ◽  
Takamasa limori ◽  
Shiro Ikegami ◽  
Shoichiro Ide ◽  
...  
Keyword(s):  
Adenylyl Cyclase ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
Inactive Analog
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