scholarly journals Coiled-coil networking shapes cell molecular machinery

2012 ◽  
Vol 23 (19) ◽  
pp. 3911-3922 ◽  
Author(s):  
Yongqiang Wang ◽  
Xinlei Zhang ◽  
Hong Zhang ◽  
Yi Lu ◽  
Haolong Huang ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Critical Role ◽  
Coiled Coil ◽  
Coiled Coils ◽  
Protein Protein Interactions ◽  
Full Spectrum ◽  
Kinetochore Assembly ◽  
Genome Wide ◽  
Molecular Machinery ◽  
Systematic Understanding

The highly abundant α-helical coiled-coil motif not only mediates crucial protein–protein interactions in the cell but is also an attractive scaffold in synthetic biology and material science and a potential target for disease intervention. Therefore a systematic understanding of the coiled-coil interactions (CCIs) at the organismal level would help unravel the full spectrum of the biological function of this interaction motif and facilitate its application in therapeutics. We report the first identified genome-wide CCI network in Saccharomyces cerevisiae, which consists of 3495 pair-wise interactions among 598 predicted coiled-coil regions. Computational analysis revealed that the CCI network is specifically and functionally organized and extensively involved in the organization of cell machinery. We further show that CCIs play a critical role in the assembly of the kinetochore, and disruption of the CCI network leads to defects in kinetochore assembly and cell division. The CCI network identified in this study is a valuable resource for systematic characterization of coiled coils in the shaping and regulation of a host of cellular machineries and provides a basis for the utilization of coiled coils as domain-based probes for network perturbation and pharmacological applications.

scholarly journals A Multiplexed Bacterial Two-Hybrid for Rapid Characterization of Protein-Protein Interactions and Iterative Protein Design

2020 ◽  
Author(s):  
W. Clifford Boldridge ◽  
Ajasja Ljubetič ◽  
Hwangbeom Kim ◽  
Nathan Lubock ◽  
Dániel Szilágyi ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Coiled Coil ◽  
Building Blocks ◽  
Coiled Coils ◽  
Next Generation ◽  
Protein Protein Interactions ◽  
Library Size ◽  
Large Sets ◽  
Synthetic Cell ◽  
Two Hybrid

AbstractMyriad biological functions require protein-protein interactions (PPIs), and engineered PPIs are crucial for applications ranging from drug design to synthetic cell circuits. Understanding and engineering specificity in PPIs is particularly challenging as subtle sequence changes can drastically alter specificity. Coiled-coils are small protein domains that have long served as a simple model for studying the sequence-determinants of specificity and have been used as modular building blocks to build large protein nanostructures and synthetic circuits. Despite their simple rules and long-time use, building large sets of well-behaved orthogonal pairs that can be used together is still challenging because predictions are often inaccurate, and, as the library size increases, it becomes difficult to test predictions at scale. To address these problems, we first developed a method called the Next-Generation Bacterial Two-Hybrid (NGB2H), which combines gene synthesis, a bacterial two-hybrid assay, and a high-throughput next-generation sequencing readout, allowing rapid exploration of interactions of programmed protein libraries in a quantitative and scalable way. After validating the NGB2H system on previously characterized libraries, we designed, built, and tested large sets of orthogonal synthetic coiled-coils. In an iterative set of experiments, we assayed more than 8,000 PPIs, used the dataset to train a novel linear model-based coiled-coil scoring algorithm, and then characterized nearly 18,000 interactions to identify the largest set of orthogonal PPIs to date with twenty-two on-target interactions.

scholarly journals High-resolution structures of a heterochiral coiled coil

2015 ◽  
Vol 112 (43) ◽  
pp. 13144-13149 ◽  
Author(s):  
David E. Mortenson ◽  
Jay D. Steinkruger ◽  
Dale F. Kreitler ◽  
Dominic V. Perroni ◽  
Gregory P. Sorenson ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Structural Information ◽  
Coiled Coil ◽  
Coiled Coils ◽  
Side Chain ◽  
Amino Acid Residues ◽  
Protein Protein Interactions ◽  
Polypeptide Chains ◽  
Packing Interactions

Interactions between polypeptide chains containing amino acid residues with opposite absolute configurations have long been a source of interest and speculation, but there is very little structural information for such heterochiral associations. The need to address this lacuna has grown in recent years because of increasing interest in the use of peptides generated from d amino acids (d peptides) as specific ligands for natural proteins, e.g., to inhibit deleterious protein–protein interactions. Coiled–coil interactions, between or among α-helices, represent the most common tertiary and quaternary packing motif in proteins. Heterochiral coiled–coil interactions were predicted over 50 years ago by Crick, and limited experimental data obtained in solution suggest that such interactions can indeed occur. To address the dearth of atomic-level structural characterization of heterochiral helix pairings, we report two independent crystal structures that elucidate coiled-coil packing between l- and d-peptide helices. Both structures resulted from racemic crystallization of a peptide corresponding to the transmembrane segment of the influenza M2 protein. Networks of canonical knobs-into-holes side-chain packing interactions are observed at each helical interface. However, the underlying patterns for these heterochiral coiled coils seem to deviate from the heptad sequence repeat that is characteristic of most homochiral analogs, with an apparent preference for a hendecad repeat pattern.

Guidelines for the assembly of novel coiled-coil structures: α-sheets and α-cylinders

2001 ◽  
Vol 68 ◽  
pp. 111-123 ◽  
Author(s):  
John Walshaw ◽  
Jennifer M. Shipway ◽  
Derek N. Woolfson
Keyword(s):  
Protein Design ◽  
Protein Interactions ◽  
Protein Structures ◽  
Coiled Coil ◽  
Data Bank ◽  
Coiled Coils ◽  
Heptad Repeat ◽  
Protein Protein Interactions ◽  
Helical Bundles ◽  
Heptad Repeats

The coiled coil is a ubiquitous motif that guides many different protein-protein interactions. The accepted hallmark of coiled coils is a seven-residue (heptad) sequence repeat. The positions of this repeat are labelled a-b-c-d-e-f-g, with residues at a and d tending to be hydrophobic. Such sequences form amphipathic α-helices, which assemble into helical bundles via knobs-into-holes interdigitation of residues from neighbouring helices. We wrote an algorithm, SOCKET, to identify this packing in protein structures, and used this to gather a database of coiled-coil structures from the Protein Data Bank. Surprisingly, in addition to commonly accepted structures with a single, contiguous heptad repeat, we identified sequences with multiple, offset heptad repeats. These 'new' sequence patterns help to explain oligomer-state specification in coiled coils. Here we focus on the structural consequences for sequences with two heptad repeats offset by two residues, i.e. a/f′-b/g′-c/a′-d/b′-e/c′-f/d′-g/e′. This sets up two hydrophobic seams on opposite sides of the helix formed. We describe how such helices may combine to bury these hydrophobic surfaces in two different ways and form two distinct structures: open 'α-sheets' and closed 'α-cylinders'. We highlight these with descriptions of natural structures and outline possibilities for protein design.

Exploring Proteomic Drug Targets, Therapeutic Strategies and Protein - Protein Interactions in Cancer: Mechanistic View

2019 ◽  
Vol 19 (6) ◽  
pp. 430-448 ◽  
Author(s):  
Khalid Bashir Dar ◽  
Aashiq Hussain Bhat ◽  
Shajrul Amin ◽  
Syed Anjum ◽  
Bilal Ahmad Reshi ◽  
...  
Keyword(s):  
Protein Interactions ◽  
High Throughput Screening ◽  
Critical Role ◽  
Cancer Therapeutics ◽  
Adaptor Proteins ◽  
Therapeutic Strategies ◽  
Small Gtpases ◽  
Beta Catenin ◽  
Protein Protein Interactions ◽  
Apoptosis Protein

Protein-Protein Interactions (PPIs) drive major signalling cascades and play critical role in cell proliferation, apoptosis, angiogenesis and trafficking. Deregulated PPIs are implicated in multiple malignancies and represent the critical targets for treating cancer. Herein, we discuss the key protein-protein interacting domains implicated in cancer notably PDZ, SH2, SH3, LIM, PTB, SAM and PH. These domains are present in numerous enzymes/kinases, growth factors, transcription factors, adaptor proteins, receptors and scaffolding proteins and thus represent essential sites for targeting cancer. This review explores the candidature of various proteins involved in cellular trafficking (small GTPases, molecular motors, matrix-degrading enzymes, integrin), transcription (p53, cMyc), signalling (membrane receptor proteins), angiogenesis (VEGFs) and apoptosis (BCL-2family), which could possibly serve as targets for developing effective anti-cancer regimen. Interactions between Ras/Raf; X-linked inhibitor of apoptosis protein (XIAP)/second mitochondria-derived activator of caspases (Smac/DIABLO); Frizzled (FRZ)/Dishevelled (DVL) protein; beta-catenin/T Cell Factor (TCF) have also been studied as prospective anticancer targets. Efficacy of diverse molecules/ drugs targeting such PPIs although evaluated in various animal models/cell lines, there is an essential need for human-based clinical trials. Therapeutic strategies like the use of biologicals, high throughput screening (HTS) and fragment-based technology could play an imperative role in designing cancer therapeutics. Moreover, bioinformatic/computational strategies based on genome sequence, protein sequence/structure and domain data could serve as competent tools for predicting PPIs. Exploring hot spots in proteomic networks represents another approach for developing targetspecific therapeutics. Overall, this review lays emphasis on a productive amalgamation of proteomics, genomics, biochemistry, and molecular dynamics for successful treatment of cancer.

scholarly journals Roles of the 14-3-3 gene family in cotton flowering

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Na Sang ◽  
Hui Liu ◽  
Bin Ma ◽  
Xianzhong Huang ◽  
Lu Zhuo ◽  
...  
Keyword(s):  
Gene Family ◽  
Protein Interactions ◽  
Leucine Zipper ◽  
Expression Patterns ◽  
Bimolecular Fluorescence Complementation ◽  
Structural Motif ◽  
Virus Induced Gene Silencing ◽  
Protein Protein Interactions ◽  
Basic Leucine Zipper ◽  
Genome Wide

Abstract Background In plants, 14-3-3 proteins, also called GENERAL REGULATORY FACTORs (GRFs), encoded by a large multigene family, are involved in protein–protein interactions and play crucial roles in various physiological processes. No genome-wide analysis of the GRF gene family has been performed in cotton, and their functions in flowering are largely unknown. Results In this study, 17, 17, 31, and 17 GRF genes were identified in Gossypium herbaceum, G. arboreum, G. hirsutum, and G. raimondii, respectively, by genome-wide analyses and were designated as GheGRFs, GaGRFs, GhGRFs, and GrGRFs, respectively. A phylogenetic analysis revealed that these proteins were divided into ε and non-ε groups. Gene structural, motif composition, synteny, and duplicated gene analyses of the identified GRF genes provided insights into the evolution of this family in cotton. GhGRF genes exhibited diverse expression patterns in different tissues. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that the GhGRFs interacted with the cotton FLOWERING LOCUS T homologue GhFT in the cytoplasm and nucleus, while they interacted with the basic leucine zipper transcription factor GhFD only in the nucleus. Virus-induced gene silencing in G. hirsutum and transgenic studies in Arabidopsis demonstrated that GhGRF3/6/9/15 repressed flowering and that GhGRF14 promoted flowering. Conclusions Here, 82 GRF genes were identified in cotton, and their gene and protein features, classification, evolution, and expression patterns were comprehensively and systematically investigated. The GhGRF3/6/9/15 interacted with GhFT and GhFD to form florigen activation complexs that inhibited flowering. However, GhGRF14 interacted with GhFT and GhFD to form florigen activation complex that promoted flowering. The results provide a foundation for further studies on the regulatory mechanisms of flowering.

scholarly journals Yeast Surface Two-hybrid for Quantitative in Vivo Detection of Protein-Protein Interactions via the Secretory Pathway

2009 ◽  
Vol 284 (24) ◽  
pp. 16369-16376 ◽  
Author(s):  
Xuebo Hu ◽  
Sungkwon Kang ◽  
Xiaoyue Chen ◽  
Charles B. Shoemaker ◽  
Moonsoo M. Jin
Keyword(s):  
Protein Interactions ◽  
Secretory Pathway ◽  
Coiled Coil ◽  
Affinity Maturation ◽  
Antibody Binding ◽  
Soluble Form ◽  
Protein Protein Interactions ◽  
Yeast Surface ◽  
Two Hybrid

A quantitative in vivo method for detecting protein-protein interactions will enhance our understanding of protein interaction networks and facilitate affinity maturation as well as designing new interaction pairs. We have developed a novel platform, dubbed “yeast surface two-hybrid (YS2H),” to enable a quantitative measurement of pairwise protein interactions via the secretory pathway by expressing one protein (bait) anchored to the cell wall and the other (prey) in soluble form. In YS2H, the prey is released either outside of the cells or remains on the cell surface by virtue of its binding to the bait. The strength of their interaction is measured by antibody binding to the epitope tag appended to the prey or direct readout of split green fluorescence protein (GFP) complementation. When two α-helices forming coiled coils were expressed as a pair of prey and bait, the amount of the prey in complex with the bait progressively decreased as the affinity changes from 100 pm to 10 μm. With GFP complementation assay, we were able to discriminate a 6-log difference in binding affinities in the range of 100 pm to 100 μm. The affinity estimated from the level of antibody binding to fusion tags was in good agreement with that measured in solution using a surface plasmon resonance technique. In contrast, the level of GFP complementation linearly increased with the on-rate of coiled coil interactions, likely because of the irreversible nature of GFP reconstitution. Furthermore, we demonstrate the use of YS2H in exploring the nature of antigen recognition by antibodies and activation allostery in integrins and in isolating heavy chain-only antibodies against botulinum neurotoxin.

scholarly journals Prioritization of genes associated with the pathogenesis of leukosis in cattle

2019 ◽  
Vol 22 (8) ◽  
pp. 1063-1069 ◽  
Author(s):  
N. S. Yudin ◽  
N. L. Podkolodnyy ◽  
T. A. Agarkova ◽  
E. V. Ignatieva
Keyword(s):  
Protein Interactions ◽  
Genome Wide Association Study ◽  
Association Studies ◽  
Mammalian Species ◽  
Genome Wide Association ◽  
Farm Animals ◽  
Genome Wide Association Studies ◽  
Protein Protein Interactions ◽  
Genome Wide ◽  
A Genome

Selection by means of genetic markers is a promising approach to the eradication of infectious diseases in farm animals, especially in the absence of effective methods of treatment and prevention. Bovine leukemia virus (BLV) is spread throughout the world and represents one of the biggest problems for the livestock production and food security in Russia. However, recent genome-wide association studies have shown that sensitivity/resistance to BLV is polygenic. The aim of this study was to create a catalog of cattle genes and genes of other mammalian species involved in the pathogenesis of BLV-induced infection and to perform gene prioritization using bioinformatics methods. Based on manually collected information from a range of open sources, a total of 446 genes were included in the catalog of cattle genes and genes of other mammals involved in the pathogenesis of BLV-induced infection. The following criteria were used to prioritize 446 genes from the catalog: (1) the gene is associated with leukemia according to a genome-wide association study; (2) the gene is associated with leukemia according to a case-control study; (3) the role of the gene in leukemia development has been studied using knockout mice; (4) protein-protein interactions exist between the gene-encoded protein and either viral particles or individual viral proteins; (5) the gene is annotated with Gene Ontology terms that are overrepresented for a given list of genes; (6) the gene participates in biological pathways from the KEGG or REACTOME databases, which are over-represented for a given list of genes; (7) the protein encoded by the gene has a high number of protein-protein interactions with proteins encoded by other genes from the catalog. Based on each criterion, a rank was assigned to each gene. Then the ranks were summarized and an overall rank was determined. Prioritization of 446 candidate genes allowed us to identify 5 genes of interest (TNF,LTB,BOLA-DQA1,BOLA-DRB3,ATF2), which can affect the sensitivity/resistance of cattle to leukemia.

scholarly journals Genome-wide protein-protein interactions and protein function exploration in cyanobacteria

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Qi Lv ◽  
Weimin Ma ◽  
Hui Liu ◽  
Jiang Li ◽  
Huan Wang ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Function ◽  
Protein Protein Interactions ◽  
Genome Wide

scholarly journals Mitochondria: Regulators of Cell Death and Survival

2002 ◽  
Vol 2 ◽  
pp. 1569-1578 ◽  
Author(s):  
David J. Granville ◽  
Roberta A. Gottlieb
Keyword(s):  
Cell Death ◽  
Conformational Changes ◽  
Protein Interactions ◽  
Apoptotic Cell ◽  
Critical Role ◽  
Death Process ◽  
Protein Protein Interactions ◽  
Ionic Changes ◽  
A Cell ◽  
Cyt C

The past 5 years has seen an intense surge in research devoted toward understanding the critical role of mitochondria in the regulation of cell death. Apoptosis can be initiated by a wide array of stimuli, inducing multiple signaling pathways that, for the most part, converge at the mitochondrion. Although classically considered the powerhouses of the cell, it is now understood that mitochondria are also “gatekeepers” that ultimately determine the fate of the cell. The mitochondrial decision as to whether a cell lives or dies is complex, involving protein-protein interactions, ionic changes, reactive oxygen species, and other mechanisms that require further elucidation. Once the death process is initiated, mitochondria undergo conformational changes, resulting in the release of cytochrome c (cyt c), caspases, endonucleases, and other factors leading to the onset and execution of apoptosis. The present review attempts to outline the complex milieu of events regulating the mitochondrial commitment to and processes involved in the implementation of the executioner phase of apoptotic cell death.

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