Plasmodium falciparum: Analysis of Protein-Protein Interactions of the 140/130/110-kDa Rhoptry Protein Complex Using Antibody and Mouse Erythrocyte Binding Assays

1993 ◽  
Vol 77 (2) ◽  
pp. 179-194 ◽  
Author(s):  
T.Y. Samyellowe
Keyword(s):  
Plasmodium Falciparum ◽  
Protein Interactions ◽  
Protein Complex ◽  
Protein Protein Interactions ◽  
Binding Assays ◽  
Mouse Erythrocyte ◽  
Erythrocyte Binding ◽  
Rhoptry Protein

scholarly journals The cytoplasmic domain of the Plasmodium falciparum ligand EBA-175 is essential for invasion but not protein trafficking

2003 ◽  
Vol 162 (2) ◽  
pp. 317-327 ◽  
Author(s):  
Tim-Wolf Gilberger ◽  
Jennifer K. Thompson ◽  
Michael B. Reed ◽  
Robert T. Good ◽  
Alan F. Cowman
Keyword(s):  
Plasmodium Falciparum ◽  
Host Cell ◽  
Protein Interactions ◽  
Protein Trafficking ◽  
Cytoplasmic Domain ◽  
Specific Protein ◽  
Host Cells ◽  
Protein Protein Interactions ◽  
Parasite Plasmodium Falciparum ◽  
Erythrocyte Binding

The invasion of host cells by the malaria parasite Plasmodium falciparum requires specific protein–protein interactions between parasite and host receptors and an intracellular translocation machinery to power the process. The transmembrane erythrocyte binding protein-175 (EBA-175) and thrombospondin-related anonymous protein (TRAP) play central roles in this process. EBA-175 binds to glycophorin A on human erythrocytes during the invasion process, linking the parasite to the surface of the host cell. In this report, we show that the cytoplasmic domain of EBA-175 encodes crucial information for its role in merozoite invasion, and that trafficking of this protein is independent of this domain. Further, we show that the cytoplasmic domain of TRAP, a protein that is not expressed in merozoites but is essential for invasion of liver cells by the sporozoite stage, can substitute for the cytoplasmic domain of EBA-175. These results show that the parasite uses the same components of its cellular machinery for invasion regardless of the host cell type and invasive form.

scholarly journals Positive selection on the Plasmodium falciparum clag2 gene encoding a component of the erythrocyte-binding rhoptry protein complex

2011 ◽  
Vol 39 (3) ◽  
pp. 77-82 ◽  
Author(s):  
Jean SF Alexandre ◽  
Morakot Kaewthamasorn ◽  
Kazuhide Yahata ◽  
Shusuke Nakazawa ◽  
Osamu Kaneko
Keyword(s):  
Plasmodium Falciparum ◽  
Positive Selection ◽  
Protein Complex ◽  
Gene Encoding ◽  
Erythrocyte Binding ◽  
Rhoptry Protein

scholarly journals An integrative approach unveils a distal encounter site for rPTPε and phospho-Src complex formation

2021 ◽  
Author(s):  
Nadendla EswarKumar ◽  
Cheng-Han Yang ◽  
Sunilkumar Tewary ◽  
Yi-Qi Yeh ◽  
Hsiao-Ching Yang ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Complex ◽  
Tyrosine Phosphatase ◽  
Complex Structure ◽  
Protein Crystallography ◽  
Md Simulations ◽  
Integrative Approach ◽  
Single Mutation ◽  
Protein Protein Interactions ◽  
Transient Nature

AbstractProtein tyrosine phosphatase: phospho-protein complex structure determination, which requires to understand how specificity is achieved at the protein level remains a significant challenge for protein crystallography and cryoEM due to the transient nature of binding interactions. Using rPTPεD1 and phospho-SrcKD as a model system, we established an integrative workflow involving protein crystallography, SAXS and pTyr-tailored MD simulations to reveal the complex formed between rPTPεD1 and phospho-SrcKD, revealing transient protein–protein interactions distal to the active site. To support our finding, we determined the associate rate between rPTPεD1 and phospho-SrcKD and showed that a single mutation on rPTPεD1 disrupts this transient interaction, resulting in the reduction of association rate and activity. Our simulations suggest that rPTPεD1 employs a binding mechanism involving conformational change prior to the engagement of cSrcKD. This integrative approach is applicable to other PTP: phospho-protein complex determination and is a general approach for elucidating transient protein surface interactions.

An activity-based probe reveals dynamic protein–protein interactions mediating IGF-1R transactivation by the GABAB receptor

2012 ◽  
Vol 443 (3) ◽  
pp. 627-634 ◽  
Author(s):  
Xin Lin ◽  
Xin Li ◽  
Ming Jiang ◽  
Linhai Chen ◽  
Chanjuan Xu ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Complex ◽  
Tyrosine Kinases ◽  
Chemical Biology ◽  
Molecular Mechanisms ◽  
Gabab Receptor ◽  
Type I ◽  
Protein Protein Interactions ◽  
Downstream Effectors ◽  
Factor Type

Many GPCRs (G-protein-coupled receptors) can activate RTKs (receptor tyrosine kinases) in the absence of RTK ligands, a phenomenon called transactivation. However, the underlying molecular mechanisms remain undefined. In the present study we investigate the molecular basis of GABAB (γ-aminobutyric acid B) receptor-mediated transactivation of IGF-1R (insulin-like growth factor type I receptor) in primary neurons. We take a chemical biology approach by developing an activity-based probe targeting the GABAB receptor. This probe enables us first to lock the GABAB receptor in an inactive state and then activate it with a positive allosteric modulator, thereby permitting monitoring of the dynamic of the protein complex associated with IGF-1R transactivation. We find that activation of the GABAB receptor induces a dynamic assembly and disassembly of a protein complex, including both receptors and their downstream effectors. FAK (focal adhesion kinase), a non-RTK, plays a key role in co-ordinating this dynamic process. Importantly, this dynamic of the GABAB receptor-associated complex is critical for transactivation and transactivation-dependent neuronal survival. The present study has identified an important mechanism underlying GPCR transactivation of RTKs, which was enabled by a new chemical biology tool generally applicable for dissecting GPCR signalling.

MLL Translocation Partners AF4, AF5q31, and ENL Associate in a Higher Order Protein Complex with CDK9 and Cyclin T1.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 99-99
Author(s):  
Min Lin ◽  
Michael L. Cleary
Keyword(s):  
Western Blotting ◽  
Protein Interactions ◽  
Protein Complex ◽  
Fusion Proteins ◽  
Higher Order ◽  
Chromosomal Translocations ◽  
Transcriptional Elongation ◽  
Protein Protein Interactions ◽  
Methyltransferase Activity ◽  
Cyclin T1

Abstract The Mixed Lineage Leukemia (MLL) gene is frequently involved in chromosomal translocations that cause acute leukemia. More than 40 different genes have been identified as MLL translocation partners, with the expression of corresponding MLL fusion proteins. The MLL protein has histone methyltransferase activity and is required for embryonic development and hematopoiesis. Several proteins have been demonstrated to associate with MLL in a macromolecular complex, which is believed to have chromatin remodeling function. However, the C-terminal SET domain of MLL, which carries the histone methyltransferase activity, is lost in all MLL fusion proteins, thus making the biochemical functions of the fusion proteins unclear. Moreover, the promiscuity of MLL translocation partners, most of them with no known functions, further complicates an understanding of MLL leukemogenic mechanisms. In this study, we purified a protein complex containing AF4, the most common MLL translocation partner, using a combination of conventional column chromatography and immunoaffinity techniques. The AF4 protein complex contains AF5q31 and ENL, two other MLL translocation partners, as well as CDK9 and Cyclin T1, a heterodimer that regulates transcriptional elongation. Gel filtration confirmed that these five proteins co-fractionate with an estimated overall size of 0.8 MDa. All protein-protein interactions were further confirmed by immunoprecipitation-western blotting from K562 cell nuclear extract. To investigate whether these protein-protein interactions are retained in corresponding MLL fusion proteins, immunoprecipitation-western blotting assays were carried out in human leukemia cell lines harboring MLL chromosomal translocations. We found that MLL-AF4, MLL-AF5q31, MLL-ENL and MLL-AF9 each associate with wild type AF4 complex components, including CDK9 and Cyclin T1. In contrast, MLL-AF6 does not associate with any of the AF4 complex components. We propose that the four nuclear MLL translocation partner proteins (AF4, AF5q31, ENL/AF9), whose translocations are found in over 75% of MLL leukemias, associate in a higher order protein complex with CDK9 and Cyclin T1 and thus function in part to regulate transcriptional elongation. The association of CDK9 and Cyclin T1 with the four MLL fusion proteins suggests a common leukemogenic mechanism that may involve transcriptional elongation, which we are currently investigating. Conversely, MLL-cytosolic fusions, e.g. MLL-AF6, appear to function independently of association with the AF4 protein complex, possibly through a homo-dimerization pathway.

scholarly journals Engineering an improved light-induced dimer (iLID) for controlling the localization and activity of signaling proteins

2014 ◽  
Vol 112 (1) ◽  
pp. 112-117 ◽  
Author(s):  
Gurkan Guntas ◽  
Ryan A. Hallett ◽  
Seth P. Zimmerman ◽  
Tishan Williams ◽  
Hayretin Yumerefendi ◽  
...  
Keyword(s):  
Binding Affinity ◽  
Protein Design ◽  
Protein Interactions ◽  
Mammalian Cell Culture ◽  
Large Change ◽  
Small Gtpase ◽  
Protein Protein Interactions ◽  
Binding Assays ◽  
Light Stimulation ◽  
Inducible Protein

The discovery of light-inducible protein–protein interactions has allowed for the spatial and temporal control of a variety of biological processes. To be effective, a photodimerizer should have several characteristics: it should show a large change in binding affinity upon light stimulation, it should not cross-react with other molecules in the cell, and it should be easily used in a variety of organisms to recruit proteins of interest to each other. To create a switch that meets these criteria we have embedded the bacterial SsrA peptide in the C-terminal helix of a naturally occurring photoswitch, the light-oxygen-voltage 2 (LOV2) domain from Avena sativa. In the dark the SsrA peptide is sterically blocked from binding its natural binding partner, SspB. When activated with blue light, the C-terminal helix of the LOV2 domain undocks from the protein, allowing the SsrA peptide to bind SspB. Without optimization, the switch exhibited a twofold change in binding affinity for SspB with light stimulation. Here, we describe the use of computational protein design, phage display, and high-throughput binding assays to create an improved light inducible dimer (iLID) that changes its affinity for SspB by over 50-fold with light stimulation. A crystal structure of iLID shows a critical interaction between the surface of the LOV2 domain and a phenylalanine engineered to more tightly pin the SsrA peptide against the LOV2 domain in the dark. We demonstrate the functional utility of the switch through light-mediated subcellular localization in mammalian cell culture and reversible control of small GTPase signaling.

scholarly journals Modified Potential Functions Result in Enhanced Predictions of a Protein Complex by All-Atom MD Simulations, Confirming a Step-wise Association Process for Native PPIs

2018 ◽  
Author(s):  
Zhen-lu Li ◽  
Matthias Buck
Keyword(s):  
Electrostatic Interaction ◽  
Protein Interactions ◽  
Protein Complex ◽  
Complex Structure ◽  
Md Simulations ◽  
Potential Functions ◽  
Amino Acid Side Chain ◽  
Native Protein ◽  
Protein Protein Interactions ◽  
Steering Mechanism

ABSTRACTNative protein-protein interactions (PPIs) are the cornerstone for understanding the structure, dynamics and mechanisms of function of protein complexes. In this study, we investigate the association of the SAM domains of the EphA2 receptor and SHIP2 enzyme by performing a combined total of 48 μs all-atom molecular dynamics (MD) simulations. While the native SAM heterodimer is only predicted at a low rate of 6.7% with the original CHARMM36 force field, the yield is increased to 16.7% and to 18.3% by scaling the vdW solute-solvent interactions (better fitting the solvation free energy of amino acid side chain analogues) and by an increase of vdW radius of guanidinium interactions, and thus a dramatic reduction of electrostatic interaction between Arg and Glu/Asn in CHARMM36m, respectively. These modifications effectively improve the overly sticky association of proteins, such as ubiquitin, using the original potential function. By analyzing the 25 native SAM complexes formed in the simulations, we find that their formation involves a pre-orientation guided by electrostatic interaction, consistent with an electrostatic steering mechanism. The complex could then transform to the native protein interaction surfaces directly from a well pre-orientated position (Δinterface-RMSD < 5Å). In other cases, modest (< 90°) orientational and/or translational adjustments are needed (5 Å <Δi-RMSD <10 Å) to the native complex. Although the tendency for non-native complexes to dissociate has nearly doubled with the modified potential functions, a re-association to the correct complex structure is still rare. Instead a most non-native complexes are undergoing configurational changes/surface searching, which do not lead to native structures on a timescale of 250 ns. These observations provide a rich picture on mechanisms of protein-protein complex formation, and suggest that computational predictions of native complex protein-protein interactions could be improved further.

Evolutionary Divergence of Plasmodium falciparum: Sequences, Protein- Protein Interactions, Pathways and Processes

2009 ◽  
Vol 9 (3) ◽  
pp. 257-271 ◽  
Author(s):  
Nidhi Tyagi ◽  
Lakshmipuram Swapna ◽  
Smita Mohanty ◽  
Garima Agarwal ◽  
Venkatraman Gowri ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Protein Interactions ◽  
Evolutionary Divergence ◽  
Protein Protein Interactions
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