scholarly journals Ancient and conserved functional interplay between Bcl-2 family proteins in the mitochondrial pathway of apoptosis

2020 ◽  
Vol 6 (40) ◽  
pp. eabc4149 ◽  
Author(s):  
Nikolay Popgeorgiev ◽  
Jaison D Sa ◽  
Lea Jabbour ◽  
Suresh Banjara ◽  
Trang Thi Minh Nguyen ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Ligand Binding ◽  
Cancer Cells ◽  
Mitochondrial Pathway ◽  
Structural Basis ◽  
Mitochondrial Outer Membrane ◽  
Chemotherapy Treatment ◽  
Trichoplax Adhaerens ◽  
Binding Groove

In metazoans, Bcl-2 family proteins are major regulators of mitochondrially mediated apoptosis; however, their evolution remains poorly understood. Here, we describe the molecular characterization of the four members of the Bcl-2 family in the most primitive metazoan, Trichoplax adhaerens. All four trBcl-2 homologs are multimotif Bcl-2 group, with trBcl-2L1 and trBcl-2L2 being highly divergent antiapoptotic Bcl-2 members, whereas trBcl-2L3 and trBcl-2L4 are homologs of proapoptotic Bax and Bak, respectively. trBax expression permeabilizes the mitochondrial outer membrane, while trBak operates as a BH3-only sensitizer repressing antiapoptotic activities of trBcl-2L1 and trBcl-2L2. The crystal structure of a trBcl-2L2:trBak BH3 complex reveals that trBcl-2L2 uses the canonical Bcl-2 ligand binding groove to sequester trBak BH3, indicating that the structural basis for apoptosis control is conserved from T. adhaerens to mammals. Finally, we demonstrate that both trBax and trBak BH3 peptides bind selectively to human Bcl-2 homologs to sensitize cancer cells to chemotherapy treatment.

scholarly journals Crystal Structure of African Swine Fever Virus A179L with the Autophagy Regulator Beclin

Viruses ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 789 ◽  
Author(s):  
Suresh Banjara ◽  
Gareth L. Shimmon ◽  
Linda K. Dixon ◽  
Christopher L. Netherton ◽  
Mark G. Hinds ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Cell Death ◽  
Ligand Binding ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
B Cell Lymphoma ◽  
Vero Cells ◽  
Fever Virus ◽  
Site Directed Mutagenesis ◽  
Binding Groove

Subversion of programmed cell death-based host defence systems is a prominent feature of infections by large DNA viruses. African swine fever virus (ASFV) is a large DNA virus and sole member of the Asfarviridae family that harbours the B-cell lymphoma 2 or Bcl-2 homolog A179L. A179L has been shown to bind to a range of cell death-inducing host proteins, including pro-apoptotic Bcl-2 proteins as well as the autophagy regulator Beclin. Here we report the crystal structure of A179L bound to the Beclin BH3 motif. A179L engages Beclin using the same canonical ligand-binding groove that is utilized to bind to pro-apoptotic Bcl-2 proteins. The mode of binding of Beclin to A179L mirrors that of Beclin binding to human Bcl-2 and Bcl-xL as well as murine γ-herpesvirus 68. The introduction of bulky hydrophobic residues into the A179L ligand-binding groove via site-directed mutagenesis ablates binding of Beclin to A179L, leading to a loss of the ability of A179L to modulate autophagosome formation in Vero cells during starvation. Our findings provide a mechanistic understanding for the potent autophagy inhibitory activity of A179L and serve as a platform for more detailed investigations into the role of autophagy during ASFV infection.

Critical residues for ligand binding in blade 2 of the propeller domain of the integrin αIIb subunit

2004 ◽  
Vol 91 (01) ◽  
pp. 111-118 ◽  
Author(s):  
Tatsushiro Tamura ◽  
Jun Yamanouchi ◽  
Shigeru Fujita ◽  
Takaaki Hato
Keyword(s):  
Crystal Structure ◽  
Ligand Binding ◽  
Binding Site ◽  
Thrombus Formation ◽  
Direct Interaction ◽  
Binding Pocket ◽  
Crystal Structure Analysis ◽  
Site Directed Mutagenesis ◽  
Structural Basis ◽  
Critical Residues

SummaryLigand binding to integrin αIIbβ3 is a key event of thrombus formation. The propeller domain of the αIIb subunit has been implicated in ligand binding. Recently, the ligand binding site of the αV propeller was determined by crystal structure analysis. However, the structural basis of ligand recognition by the αIIb propeller remains to be determined. In this study, we conducted site-directed mutagenesis of all residues located in the loops extending above blades 2 and 4 of the αIIb propeller, which are spatially close to, but distinct from, the loops that contain the binding site for an RGD ligand in the crystal structure of the αV propeller. Replacement by alanine of Q111, H112 or N114 in the loop within the blade 2 (the W2:2-3 loop in the propeller model) abolished binding of a ligand-mimetic antibody and fibrinogen to αIIbβ3 induced by different types of integrin activation including activation of αIIbβ3 by β3 cytoplasmic mutation. CHO cells stably expressing recombinant αIIbβ3 bearing Q111A, H112A or N114A mutation did not exhibit αIIbβ3mediated adhesion to fibrinogen. According to the crystal structure of αVβ3, the αV residue corresponding to αIIbN114 is exposed on the integrin surface and close to the RGD binding site. These results suggest that the Q111, H112 and N114 residues in the loop within blade 2 of the αIIb propeller are critical for ligand binding, possibly because of direct interaction with ligands or modulation of the RGD binding pocket.

Crystal structures of a β-trefoil lectin from Entamoeba histolytica in monomeric and a novel disulfide bond-mediated dimeric forms

Glycobiology ◽  
2020 ◽  
Vol 30 (7) ◽  
pp. 474-488 ◽  
Author(s):  
Farha Khan ◽  
Devanshu Kurre ◽  
K Suguna
Keyword(s):  
Crystal Structure ◽  
Crystal Structures ◽  
Entamoeba Histolytica ◽  
Disulfide Bond ◽  
Structural Basis ◽  
Free Form ◽  
Biophysical Characterization ◽  
Biophysical Studies ◽  
First Time

Abstract β-Trefoil lectins are galactose/N-acetyl galactosamine specific lectins, which are widely distributed across all kingdoms of life and are known to perform several important functions. However, there is no report available on the characterization of these lectins from protozoans. We have performed structural and biophysical studies on a β-trefoil lectin from Entamoeba histolytica (EntTref), which exists as a mixture of monomers and dimers in solution. Further, we have determined the affinities of EntTref for rhamnose, galactose and different galactose-linked sugars. We obtained the crystal structure of EntTref in a sugar-free form (EntTref_apo) and a rhamnose-bound form (EntTref_rham). A novel Cys residue-mediated dimerization was revealed in the crystal structure of EntTref_apo while the structure of EntTref_rham provided the structural basis for the recognition of rhamnose by a β-trefoil lectin for the first time. To the best of our knowledge, this is the only report of the structural, functional and biophysical characterization of a β-trefoil lectin from a protozoan source and the first report of Cys-mediated dimerization in this class of lectins.

Structural basis ofDeerpox virus-mediated inhibition of apoptosis

2015 ◽  
Vol 71 (8) ◽  
pp. 1593-1603 ◽  
Author(s):  
Denis R. Burton ◽  
Sofia Caria ◽  
Bevan Marshall ◽  
Michele Barry ◽  
Marc Kvansakul
Keyword(s):  
Structural Analysis ◽  
Ligand Binding ◽  
Structural Basis ◽  
Inhibitor Of Apoptosis ◽  
Defence Mechanism ◽  
Partial Obstruction ◽  
Host Cell Apoptosis ◽  
Infected Cells ◽  
Binding Groove ◽  
Innate Defence

Apoptosis is a key innate defence mechanism to eliminate virally infected cells. To counteract premature host-cell apoptosis, poxviruses have evolved numerous molecular strategies, including the use of Bcl-2 proteins, to ensure their own survival. Here, it is reported that theDeerpox virusinhibitor of apoptosis, DPV022, only engages a highly restricted set of death-inducing Bcl-2 proteins, including Bim, Bax and Bak, with modest affinities. Structural analysis reveals that DPV022 adopts a Bcl-2 fold with a dimeric domain-swapped topology and binds pro-death Bcl-2 proteinsviatwo conserved ligand-binding grooves found on opposite sides of the dimer. Structures of DPV022 bound to Bim, Bak and Bax BH3 domains reveal that a partial obstruction of the binding groove is likely to be responsible for the modest affinities of DPV022 for BH3 domains. These findings reveal that domain-swapped dimeric Bcl-2 folds are not unusual and may be found more widely in viruses. Furthermore, the modest affinities of DPV022 for pro-death Bcl-2 proteins suggest that two distinct classes of anti-apoptotic viral Bcl-2 proteins exist: those that are monomeric and tightly bind a range of death-inducing Bcl-2 proteins, and others such as DPV022 that are dimeric and only bind a very limited number of death-inducing Bcl-2 proteins with modest affinities.

scholarly journals Crystal Structure of African Swine Fever Virus A179L with the Autophagy Regulator Beclin

2019 ◽  
Author(s):  
Suresh Banjara ◽  
Gareth Shimmon ◽  
Linda Dixon ◽  
Christopher Netherton ◽  
Mark Hinds ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Cell Death ◽  
Ligand Binding ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
B Cell Lymphoma ◽  
Vero Cells ◽  
Fever Virus ◽  
Site Directed Mutagenesis ◽  
Binding Groove

Subversion of programmed cell death-based host defence systems is a prominent feature of infections by large DNA viruses. African swine fever virus (ASFV) is a large DNA virus and sole member of the Asfarviridae family that harbors the B-cell lymphoma 2 or Bcl-2 homolog A179L. A179L has been shown to bind to a range of cell death inducing host proteins including pro-apoptotic Bcl-2 proteins as well as the autophagy regulator Beclin. Here we report the crystal structure of A179L bound to the Beclin BH3 motif. A179L engages Beclin using the same canonical ligand binding groove that is utilized to bind to pro-apoptotic Bcl-2 proteins. The mode of binding of Beclin to A179L mirrors that of Beclin binding to human Bcl-2 and Bcl-xL as well as murine gamma-herpesvirus 68. Introduction of bulky hydrophobic residues into the A179L ligand binding groove via site directed mutagenesis ablates binding of Beclin to A179L, leading to a loss of ability of A179L to modulate autophagosome formation in Vero cells during starvation. Our findings provide a mechanistic understanding for the potent autophagy inhibitory activity of A179L and serve as a platform for more detailed investigations into the role of autophagy during ASFV infection.

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