Halophilic Adaptation:  Novel Solvent Protein Interactions Observed in the 2.9 and 2.6 Å Resolution Structures of the Wild Type and a Mutant of Malate Dehydrogenase fromHaloarcula marismortui‡

Several studies have indicated that the central pair of microtubules and their associated structures play a significant role in regulating flagellar motility. To begin a molecular analysis of these components we have generated central apparatus-defective mutants in Chlamydomonas reinhardtii using insertional mutagenesis. One paralyzed mutant recovered in our screen, D2, is an allele of a previously identified mutant, pf16. Mutant cells have paralyzed flagella, and the C1 microtubule of the central apparatus is missing in isolated axonemes. We have cloned the wild-type PF16 gene and confirmed its identity by rescuing pf16 mutants upon transformation. The rescued pf16 cells were wild-type in motility and in axonemal ultrastructure. A full-length cDNA clone for PF16 was obtained and sequenced. Database searches using the predicted 566 amino acid sequence of PF16 indicate that the protein contains eight contiguous armadillo repeats. A number of proteins with diverse cellular functions also contain armadillo repeats including pendulin, Rch1, importin, SRP-1, and armadillo. An antibody was raised against a fusion protein expressed from the cloned cDNA. Immunofluorescence labeling of wild-type flagella indicates that the PF16 protein is localized along the length of the flagella while immunogold labeling further localizes the PF16 protein to a single microtubule of the central pair. Based on the localization results and the presence of the armadillo repeats in this protein, we suggest that the PF16 gene product is involved in protein-protein interactions important for C1 central microtubule stability and flagellar motility.

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Extracting Biological Significant Subnetworks from Protein-Protein Interactions Induced by Differentially Expressed Genes of HIV-1 Vpr Variants

International Journal of System Dynamics Applications ◽

10.4018/ijsda.2015100103 ◽

2015 ◽

Vol 4 (4) ◽

pp. 35-51 ◽

Cited By ~ 1

Author(s):

Bandana Barman ◽

Anirban Mukhopadhyay

Keyword(s):

Differentially Expressed Genes ◽

Protein Interaction ◽

Protein Interactions ◽

Protein Interaction Network ◽

Interaction Network ◽

Differentially Expressed ◽

Wild Type ◽

Protein Protein Interaction ◽

Ppi Networks ◽

Hiv 1

Identification of protein interaction network is very important to find the cell signaling pathway for a particular disease. The authors have found the differentially expressed genes between two sample groups of HIV-1. Samples are wild type HIV-1 Vpr and HIV-1 mutant Vpr. They did statistical t-test and found false discovery rate (FDR) to identify the genes increased in expression (up-regulated) or decreased in expression (down-regulated). In the test, the authors have computed q-values of test to identify minimum FDR which occurs. As a result they found 172 differentially expressed genes between their sample wild type HIV-1 Vpr and HIV-1 mutant Vpr, R80A. They found 68 up-regulated genes and 104 down-regulated genes. From the 172 differentially expressed genes the authors found protein-protein interaction network with string-db and then clustered (subnetworks) the PPI networks with cytoscape3.0. Lastly, the authors studied significance of subnetworks with performing gene ontology and also studied the KEGG pathway of those subnetworks.

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An innovative method for the selection of inhibitors of the viral spike-glycoprotein of the SARS-CoV

Journal of Bioinformatics and Computational Biology ◽

10.1142/s0219720020500432 ◽

2020 ◽

pp. 2050043

Author(s):

T. V. Koshlan ◽

K. G. Kulikov

Keyword(s):

Active Center ◽

Protein Interactions ◽

Software Package ◽

Wild Type ◽

Spike Glycoprotein ◽

Innovative Method ◽

Detailed Method ◽

Step Algorithm ◽

Natural Peptides ◽

Selection Of

This paper has developed and described a detailed method for selecting inhibitors based on modified natural peptides for the SARS-CoV BJ01 spike-glycoprotein. The selection of inhibitors is carried out by increasing the affinity of the peptide to the active center of the protein. This paper also provides a step-by-step algorithm for analyzing the affinity of protein interactions and presents an analysis of energy interactions between the active center of a protein and the wild-type peptide interacting with it, taking into account modifications of the latter. A description of the software package that implements the presented algorithm is given on the website https://binomlabs.com/covid19 .

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ATXN1 N-terminal region explains the binding differences of wild-type and expanded forms

BMC Medical Genomics ◽

10.1186/s12920-019-0594-4 ◽

2019 ◽

Vol 12 (1) ◽

Author(s):

Sara Rocha ◽

Jorge Vieira ◽

Noé Vázquez ◽

Hugo López-Fernández ◽

Florentino Fdez-Riverola ◽

...

Keyword(s):

Protein Interactions ◽

Protein Structures ◽

Interaction Strength ◽

Terminal Region ◽

Spinocerebellar Ataxia Type ◽

Wild Type ◽

Binding Interface ◽

Strength Of Interaction ◽

The Difference ◽

Polyq Expansion

Abstract Background Wild-type (wt) polyglutamine (polyQ) regions are implicated in stabilization of protein-protein interactions (PPI). Pathological polyQ expansion, such as that in human Ataxin-1 (ATXN1), that causes spinocerebellar ataxia type 1 (SCA1), results in abnormal PPI. For ATXN1 a larger number of interactors has been reported for the expanded (82Q) than the wt (29Q) protein. Methods To understand how the expanded polyQ affects PPI, protein structures were predicted for wt and expanded ATXN1, as well as, for 71 ATXN1 interactors. Then, the binding surfaces of wt and expanded ATXN1 with the reported interactors were inferred. Results Our data supports that the polyQ expansion alters the ATXN1 conformation and that it enhances the strength of interaction with ATXN1 partners. For both ATXN1 variants, the number of residues at the predicted binding interface are greater after the polyQ, mainly due to the AXH domain. Moreover, the difference in the interaction strength of the ATXN1 variants was due to an increase in the number of interactions at the N-terminal region, before the polyQ, for the expanded form. Conclusions There are three regions at the AXH domain that are essential for ATXN1 PPI. The N-terminal region is responsible for the strength of the PPI with the ATXN1 variants. How the predicted motifs in this region affect PPI is discussed, in the context of ATXN1 post-transcriptional modifications.

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The Effect of Proline cis-trans Isomerization on the Folding of the C-Terminal SH2 Domain from p85

International Journal of Molecular Sciences ◽

10.3390/ijms21010125 ◽

2019 ◽

Vol 21 (1) ◽

pp. 125

Author(s):

Francesca Troilo ◽

Francesca Malagrinò ◽

Lorenzo Visconti ◽

Angelo Toto ◽

Stefano Gianni

Keyword(s):

Protein Interactions ◽

Specific Interaction ◽

Sh2 Domain ◽

Regulatory Subunit ◽

Wild Type ◽

Protein Protein Interactions ◽

Sh2 Domains ◽

Trans Isomerization ◽

A Site ◽

Kinetics Of

SH2 domains are protein domains that modulate protein–protein interactions through a specific interaction with sequences containing phosphorylated tyrosines. In this work, we analyze the folding pathway of the C-terminal SH2 domain of the p85 regulatory subunit of the protein PI3K, which presents a proline residue in a cis configuration in the loop between the βE and βF strands. By employing single and double jump folding and unfolding experiments, we demonstrate the presence of an on-pathway intermediate that transiently accumulates during (un)folding. By comparing the kinetics of folding of the wild-type protein to that of a site-directed variant of C-SH2 in which the proline was replaced with an alanine, we demonstrate that this intermediate is dictated by the peptidyl prolyl cis-trans isomerization. The results are discussed in the light of previous work on the effect of peptidyl prolyl cis-trans isomerization on folding events.

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Genetic Manipulation of Key Photosynthetic Enzymes in the C4 Plant Flaveria bidentis

Functional Plant Biology ◽

10.1071/pp97028 ◽

1997 ◽

Vol 24 (4) ◽

pp. 477 ◽

Cited By ~ 50

Author(s):

Robert T. Furbank ◽

Julie A. Chitty ◽

Colin L.D. Jenkins ◽

William C. Taylor ◽

Stephen J. Trevanion ◽

...

Keyword(s):

Enzyme Activity ◽

Malate Dehydrogenase ◽

Genetic Manipulation ◽

Photosynthetic Performance ◽

Covalent Modification ◽

Wild Type ◽

Bisphosphate Carboxylase ◽

Photosynthetic Enzymes ◽

Flaveria Bidentis

The NADP-malic enzyme type C4 dicot Flaveria bidentis (L.) Kuntze was transformed with antisense and cosense gene constructs that resulted in specific decreases in single photosynthetic enzymes. The enzymes targeted were ribulose-1,5-bisphosphate carboxylase/oxygenase [EC 4.1.1.39] (Rubisco), pyruvate, Pi dikinase [EC 2.7.9.1] (PPDK) and NADP malate dehydrogenase [EC 1.1.1.82] (NADP-MDH). These enzymes were chosen as targets because they have low activity compared to photosynthetic rates (Rubisco), are subject to complex covalent regulation (NADP-MDH), or both (PPDK). T1 progeny of a number of lines of these transformants were examined for the effects of these gene constructs on enzyme levels and photosynthetic performance. Rubisco antisense transformants expressing between 15 and 100% of wild-type enzyme activity were obtained. Pyruvate, Pi dikinase antisense lines were obtained with 40–100% wild-type levels. NADP malate dehydrogenase sense constructs caused a co-suppression of enzyme activity with some lines containing less than 2% of wild- type activity. Under saturating illumination, the control coefficients for these enzymes were determined to be up to 0.7 for Rubisco, 0.2–0.3 for PPDK and effectively zero for NADP-MDH. The implications of these observations for the regulation of photosynthetic flux and metabolism in C4 plants and the role of regulation by covalent modification are discussed.

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Proximal Protein Interaction Landscape of RAS Paralogs

Cancers ◽

10.3390/cancers12113326 ◽

2020 ◽

Vol 12 (11) ◽

pp. 3326 ◽

Cited By ~ 1

Author(s):

Benoît Béganton ◽

Etienne Coyaud ◽

Estelle M. N. Laurent ◽

Alain Mangé ◽

Julien Jacquemetton ◽

...

Keyword(s):

Protein Interaction ◽

Protein Interactions ◽

Biological Activities ◽

Mammalian Species ◽

Variable Region ◽

Specific Protein ◽

Protein Networks ◽

Ras Proteins ◽

Wild Type ◽

Protein Protein Interaction

RAS proteins (KRAS, NRAS and HRAS) are frequently activated in different cancer types (e.g., non-small cell lung cancer, colorectal cancer, melanoma and bladder cancer). For many years, their activities were considered redundant due to their high degree of sequence homology (80% identity) and their shared upstream and downstream protein partners. However, the high conservation of the Hyper-Variable-Region across mammalian species, the preferential activation of different RAS proteins in specific tumor types and the specific post-translational modifications and plasma membrane-localization of each paralog suggest they could ensure discrete functions. To gain insights into RAS proteins specificities, we explored their proximal protein–protein interaction landscapes using the proximity-dependent biotin identification technology (BioID) in Flp-In T-REx 293 cell lines stably transfected and inducibly expressing wild type KRAS4B, NRAS or HRAS. We identified more than 800 high-confidence proximal interactors, allowing us to propose an unprecedented comparative analysis of wild type RAS paralogs protein networks. These data bring novel information on poorly characterized RAS functions, e.g., its putative involvement in metabolic pathways, and on shared as well as paralog-specific protein networks that could partially explain the complexity of RAS functions. These networks of protein interactions open numerous avenues to better understand RAS paralogs biological activities.

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LMP-1's Transmembrane Domains Encode Multiple Functions Required for LMP-1's Efficient Signaling

Journal of Virology ◽

10.1128/jvi.76.22.11551-11560.2002 ◽

2002 ◽

Vol 76 (22) ◽

pp. 11551-11560 ◽

Cited By ~ 18

Author(s):

Ajamete Kaykas ◽

Kathleen Worringer ◽

Bill Sugden

Keyword(s):

Protein Interactions ◽

Tumor Necrosis Factor Receptor ◽

Wild Type ◽

Multiple Functions ◽

Barr Virus ◽

Signaling Domain ◽

Epstein Barr ◽

Membrane Spanning ◽

Carboxy Terminal ◽

Membrane Spanning Domains

ABSTRACT The latent membrane protein-1 (LMP-1) of Epstein-Barr virus (EBV) contributes to the proliferation of infected B lymphocytes by signaling through its binding to cellular signaling molecules. It apparently mimics members of the tumor necrosis factor receptor family, in particular, CD40, by binding a similar set of cellular molecules as does CD40. LMP-1 differs dramatically in its structure from CD40. LMP-1 has six membrane-spanning domains as opposed to CD40's one. LMP-1 also differs from CD40 in its apparent independence of a ligand for its signaling. We have examined the role of LMP-1's membrane-spanning domains in its signaling. Their substitution with six membrane-spanning domains from the LMP-2A protein of EBV yields a derivative which neither coimmunoprecipitates with LMP-1 nor signals to increase the activity of NF-κB as does wild-type LMP-1. These observations indicate that LMP-1 has specific sequences in its membrane-spanning domains required for these activities. LMP-1's first and sixth membrane-spanning domains have multiple leucine residues potentially similar to leucine-heptad motifs that can mediate protein-protein interactions in membranes (Gurezka et al., J. Biol. Chem. 274:9265-9270, 1999). Substitution of seven leucines in LMP-1's sixth membrane-spanning domain has no effect on its function, whereas similar substitutions in its first membrane-spanning domain yielded a derivative which aggregates as does wild-type LMP-1 but has only 3% of wild-type's ability to signal through NF-κB. Importantly, this derivative complements a mutant of LMP-1 with wild-type membrane-spanning domains but no carboxy-terminal signaling domain. These findings together indicate that the membrane-spanning domains of LMP-1 contribute multiple functions to its signaling.

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Hemophilia A mutations associated with 1-stage/2-stage activity discrepancy disrupt protein-protein interactions within the triplicated A domains of thrombin-activated factor VIIIa

Blood ◽

10.1182/blood.v97.3.685 ◽

2001 ◽

Vol 97 (3) ◽

pp. 685-691 ◽

Cited By ~ 62

Author(s):

Steven W. Pipe ◽

Evgueni L. Saenko ◽

Angela N. Eickhorst ◽

Geoffrey Kemball-Cook ◽

Randal J. Kaufman

Keyword(s):

Protein Interactions ◽

Hemophilia A ◽

Specific Activity ◽

Homology Model ◽

Optical Biosensor ◽

Hydrophobic Core ◽

Wild Type ◽

Activity Assay ◽

Subunit Dissociation ◽

A Domains

Abstract Thrombin-activated factor VIII (FVIIIa) is a heterotrimer with the A2 subunit (amino acid residues 373-740) in a weak ionic interaction with the A1 and A3-C1-C2 subunits. Dissociation of the A2 subunit correlates with inactivation of FVIIIa. Patients with hemophilia A have been described whose plasmas display a discrepancy between their FVIII activities, where the 1-stage activity assay displays greater activity than the 2-stage activity assay. The molecular basis for one of these mutations, ARG531HIS, is an increased rate of A2 subunit dissociation. Examination of a homology model of the A domains of FVIII predicted ARG531 to lie at the interface of the A1 and A2 subunits and stabilize their interaction. Indeed, patients with mutations either directly contacting ARG531 (ALA284GLU, ALA284PRO) or closely adjacent to the A1-A2 interface in the tightly packed hydrophobic core (SER289LEU) have the same phenotype of 1-stage/2-stage discrepancy. TheALA284GLU andSER289LEU mutations in FVIII were produced by transfection of COS-1 monkey cells. Compared to FVIII wild-type both mutants had reduced specific activity by 1-stage clotting activity and at least a 2-fold lower activity by 2-stage analysis (COAMATIC), similar to the reported clinical data. Analysis of immunoaffinity purified ALA284GLU andSER289LEU proteins in an optical biosensor demonstrated that A2 dissociation was 3-fold faster for both FVIIIa mutants compared to FVIIIa wild-type. Therefore, these mutations within the A1 subunit of FVIIIa introduce a similar destabilization of the FVIIIa heterotrimer compared to the ARG531HISmutation within the A2 subunit and support that these residues stabilize the A domain interface of FVIIIa.

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