scholarly journals Identification of critical residues in the colicin E9 DNase binding region of the Im9 protein

1997 ◽  
Vol 323 (3) ◽  
pp. 823-831 ◽  
Author(s):  
Michael J. OSBORNE ◽  
Russell WALLIS ◽  
Kit-Yi LEUNG ◽  
Glyn WILLIAMS ◽  
Lu-Yun LIAN ◽  
...  
Keyword(s):  
Binding Site ◽  
Protein Complexes ◽  
Dissociation Rate ◽  
15N Nmr ◽  
Binding Region ◽  
Nmr Studies ◽  
Immunity Protein ◽  
Wide Range ◽  
Protein Dissociation ◽  
Colicin E9

1H–15N NMR studies, in conjunction with mutagenesis experiments, have been used to delineate the DNase-binding surface of the colicin E9 inhibitor protein Im9 (where Im stands for immunity protein). Complexes were formed between the 15 kDa unlabelled E9 DNase domain and the 9.5 kDa Im9 protein uniformly labelled with 15N. Approx. 90% of the amide resonances of the bound Im9 were assigned and spectral parameters obtained from 1H–15N heteronuclear single quantum coherence (HSQC) spectra were compared with those for the free Im9 assigned previously. Many of the amide resonances were shifted on complex formation, some by more than 2 p.p.m. in the 15N dimension and more than 0.5 p.p.m. in the 1H dimension. Most of the strongly shifted amides are located on the surfaces of two of the four helices, helix II and helix III. Whereas helix II had already been identified through genetic and biochemical investigations as an important determinant of biological specificity, helix III had not previously been implicated in binding to the DNase. To test the robustness of the NMR-delineated DNase-binding site, a selection of Im9 alanine mutants were constructed and their dissociation rate constants from E9 DNase-immunity protein complexes quantified by radioactive subunit exchange kinetics. Their off-rates correlated well with the NMR perturbation analysis; for example, residues that were highly perturbed in HSQC experiments, such as residues 34 (helix II) and 54 (helix III), had a marked effect on the DNase–immunity protein dissociation rate when replaced by alanine. The NMR and mutagenesis data are consistent with a DNase-binding region on Im9 composed of invariant residues in helix III and variable residues in helix II. The relationship of this binding site model to the wide range of affinities (Kd values in the range 10-4 to 10-16 M) that have been measured for cognate and non-cognate colicin DNase–immunity protein interactions is discussed.

Clusters in an Intrinsically Disordered Protein Create a Protein-Binding Site:  The TolB-Binding Region of Colicin E9†

Biochemistry ◽  
2005 ◽  
Vol 44 (34) ◽  
pp. 11496-11507 ◽  
Author(s):  
Kaeko Tozawa ◽  
Colin J. Macdonald ◽  
Christopher N. Penfold ◽  
Richard James ◽  
Colin Kleanthous ◽  
...  
Keyword(s):  
Protein Binding ◽  
Binding Site ◽  
Intrinsically Disordered Protein ◽  
Protein Binding Site ◽  
Binding Region ◽  
Intrinsically Disordered ◽  
Disordered Protein ◽  
Colicin E9

Protein-Protein Interactions in Colicin E9 DNase-Immunity Protein Complexes. 2. Cognate and Noncognate Interactions That Span the Millilmolar to Femptomolar Affinity Range

Biochemistry ◽  
1995 ◽  
Vol 34 (42) ◽  
pp. 13751-13759 ◽  
Author(s):  
Russell Wallis ◽  
Kit-Yi Leung ◽  
Ansgar J. Pommer ◽  
Hortense Videler ◽  
Geoffrey R. Moore ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Complexes ◽  
Protein Protein Interactions ◽  
Immunity Protein ◽  
Colicin E9

scholarly journals Computational studies of protein–protein dissociation by statistical potential and coarse-grained simulations: a case study on interactions between colicin E9 endonuclease and immunity proteins

2019 ◽  
Vol 21 (5) ◽  
pp. 2463-2471 ◽  
Author(s):  
Zhaoqian Su ◽  
Yinghao Wu
Keyword(s):  
Protein Complexes ◽  
Simulation Method ◽  
Coarse Grained ◽  
Computational Studies ◽  
Knowledge Based ◽  
Protein Dissociation ◽  
Coarse Grained Simulations ◽  
Dissociation Mechanisms ◽  
Colicin E9

A coarse-grained simulation method and a knowledge-based potential were developed to explore the dissociation mechanisms of protein complexes.

Localization of a Vitronectin Binding Region of Plasminogen Activator Inhibitor-1

1995 ◽  
Vol 73 (05) ◽  
pp. 829-834 ◽  
Author(s):  
Jaya Padmanabhan ◽  
David C Sane
Keyword(s):  
Binding Site ◽  
Plasminogen Activator ◽  
Inhibitory Activity ◽  
Plasminogen Activator Inhibitor ◽  
Structural Homology ◽  
Binding Region ◽  
Plasminogen Activator Inhibitor 1 ◽  
Activator Inhibitor ◽  
Pai 1

SummaryThe PAI-1 binding site for VN was studied using two independent methods. PAI-1 was cleaved by Staph V8 protease, producing 8 fragments, only 2 of which bound to [125I]-VN. These fragments were predicted to overlap between residues 91-130. Since PAI-2 has structural homology to PAI-1, but does not bind to vitronectin, chimeras of PAI-1 and PAI-2 were constructed. Four chimeras, containing PAI-1 residues 1-70,1-105,1-114, and 1-167 were constructed and expressed in vitro. PAI-1, PAI-2, and all of the chimeras retained inhibitory activity for t-PA, but only the chimera containing PAI-1 residues 1-167 formed a complex with VN. Together, these results predict that the VN binding site of PAI-1 is between residues 115-130.

Chaperonin as the normal controls and pathological anti-stress response in the human reproductive system

2016 ◽  
pp. 126-129
Author(s):  
M. Makarenko ◽  
◽  
D. Hovsyeyev ◽  
L. Sydoryk ◽  
◽  
...  
Keyword(s):  
Reproductive System ◽  
Stress Proteins ◽  
Physiological Stress ◽  
Protein Complexes ◽  
Reproductive Function ◽  
Intercellular Signaling ◽  
Toll Like Receptors ◽  
Wide Range ◽  
Protein Functions ◽  
And Function

Different kinds of physiological stress cause mass changes in the cells, including the changes in the structure and function of the protein complexes and in separate molecules. The protein functions is determined by its folding (the spatial conclusion), which depends on the functioning of proteins of thermal shock- molecular chaperons (HSPs) or depends on the stress proteins, that are high-conservative; specialized proteins that are responsible for the correct proteinaceous folding. The family of the molecular chaperones/ chaperonins/ Hsp60 has a special place due to the its unique properties of activating the signaling cascades through the system of Toll-like receptors; it also stimulates the cells to produce anti- inflammatory cytokines, defensins, molecules of cell adhesion and the molecules of MHC; it functions as the intercellular signaling molecule. The pathological role of Hsp60 is established in a wide range of illnesses, from diabetes to atherosclerosis, where Hsp60 takes part in the regulation of both apoptosis and the autoimmune processes. The presence of the HSPs was found in different tissues that are related to the reproductive system. Key words: molecular chaperons (HSPs), Toll-like receptors, reproductive function, natural auto antibody.

15N NMR studies provide insights into physico-chemical properties of room-temperature ionic liquids

2021 ◽  
Author(s):  
Christoph Wiedemann ◽  
David Fushman ◽  
Frank Bordusa
Keyword(s):  
Ionic Liquids ◽  
Room Temperature ◽  
Chemical Properties ◽  
Room Temperature Ionic Liquids ◽  
15N Nmr ◽  
Nmr Studies ◽  
Physico Chemical ◽  
Alternative Solvents

Ionic liquids (ILs) have gained a lot of attention as alternative solvents in many fields of science in the last two decades. It is known that the type of anion...

scholarly journals The Structure of the Cysteine-Rich Domain of Plasmodium falciparum P113 Identifies the Location of the RH5 Binding Site

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Ivan Campeotto ◽  
Francis Galaway ◽  
Shahid Mehmood ◽  
Lea K. Barfod ◽  
Doris Quinkert ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Binding Site ◽  
Structural Information ◽  
Blood Stage ◽  
Site Directed Mutagenesis ◽  
Effective Vaccine ◽  
Binding Region ◽  
Fab Fragment ◽  
Content Type ◽  
Blood Stage Malaria

ABSTRACT Plasmodium falciparum RH5 is a secreted parasite ligand that is essential for erythrocyte invasion through direct interaction with the host erythrocyte receptor basigin. RH5 forms a tripartite complex with two other secreted parasite proteins, CyRPA and RIPR, and is tethered to the surface of the parasite through membrane-anchored P113. Antibodies against RH5, CyRPA, and RIPR can inhibit parasite invasion, suggesting that vaccines containing these three components have the potential to prevent blood-stage malaria. To further explore the role of the P113-RH5 interaction, we selected monoclonal antibodies against P113 that were either inhibitory or noninhibitory for RH5 binding. Using a Fab fragment as a crystallization chaperone, we determined the crystal structure of the RH5 binding region of P113 and showed that it is composed of two domains with structural similarities to rhamnose-binding lectins. We identified the RH5 binding site on P113 by using a combination of hydrogen-deuterium exchange mass spectrometry and site-directed mutagenesis. We found that a monoclonal antibody to P113 that bound to this interface and inhibited the RH5-P113 interaction did not inhibit parasite blood-stage growth. These findings provide further structural information on the protein interactions of RH5 and will be helpful in guiding the development of blood-stage malaria vaccines that target RH5. IMPORTANCE Malaria is a deadly infectious disease primarily caused by the parasite Plasmodium falciparum. It remains a major global health problem, and there is no highly effective vaccine. A parasite protein called RH5 is centrally involved in the invasion of host red blood cells, making it—and the other parasite proteins it interacts with—promising vaccine targets. We recently identified a protein called P113 that binds RH5, suggesting that it anchors RH5 to the parasite surface. In this paper, we use structural biology to locate and characterize the RH5 binding region on P113. These findings will be important to guide the development of new antimalarial vaccines to ultimately prevent this disease, which affects some of the poorest people on the planet.

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