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Current Protein & Peptide Science 2020Proteins are the most critical executive molecules by responding to the instructions stored in the genetic materials in any form of life. More frequently, proteins do... (Review)
Review
Proteins are the most critical executive molecules by responding to the instructions stored in the genetic materials in any form of life. More frequently, proteins do their jobs by acting as a roleplayer that interacts with other protein(s), which is more evident when the function of a protein is examined in the real context of a cell. Identifying the interactions between (or amongst) proteins is very crucial for the biochemistry investigation of an individual protein and for the attempts aiming to draw a holo-picture for the interacting members at the scale of proteomics (or protein-protein interactions mapping). Here, we introduced the currently available reporting systems that can be used to probe the interaction between candidate protein pairs based on the fragment complementation of some particular proteins. Emphasis was put on the principles and details of experimental design. These systems are dihydrofolate reductase (DHFR), β-lactamase, tobacco etch virus (TEV) protease, luciferase, β- galactosidase, GAL4, horseradish peroxidase (HRP), focal adhesion kinase (FAK), green fluorescent protein (GFP), and ubiquitin.
Topics: Animals; Binding Sites; Biological Assay; DNA-Binding Proteins; Endopeptidases; Escherichia coli; Focal Adhesion Protein-Tyrosine Kinases; Green Fluorescent Proteins; Horseradish Peroxidase; Luciferases; Peptide Fragments; Potyvirus; Protein Binding; Protein Interaction Mapping; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Tetrahydrofolate Dehydrogenase; Transcription Factors; Ubiquitin; beta-Galactosidase; beta-Lactamases
PubMed: 32053071
DOI: 10.2174/1389203721666200213102829 -
Nature May 2023Peroxisomes are organelles that carry out β-oxidation of fatty acids and amino acids. Both rare and prevalent diseases are caused by their dysfunction. Among...
Peroxisomes are organelles that carry out β-oxidation of fatty acids and amino acids. Both rare and prevalent diseases are caused by their dysfunction. Among disease-causing variant genes are those required for protein transport into peroxisomes. The peroxisomal protein import machinery, which also shares similarities with chloroplasts, is unique in transporting folded and large, up to 10 nm in diameter, protein complexes into peroxisomes. Current models postulate a large pore formed by transmembrane proteins; however, so far, no pore structure has been observed. In the budding yeast Saccharomyces cerevisiae, the minimum transport machinery includes the membrane proteins Pex13 and Pex14 and the cargo-protein-binding transport receptor, Pex5. Here we show that Pex13 undergoes liquid-liquid phase separation (LLPS) with Pex5-cargo. Intrinsically disordered regions in Pex13 and Pex5 resemble those found in nuclear pore complex proteins. Peroxisomal protein import depends on both the number and pattern of aromatic residues in these intrinsically disordered regions, consistent with their roles as 'stickers' in associative polymer models of LLPS. Finally, imaging fluorescence cross-correlation spectroscopy shows that cargo import correlates with transient focusing of GFP-Pex13 and GFP-Pex14 on the peroxisome membrane. Pex13 and Pex14 form foci in distinct time frames, suggesting that they may form channels at different saturating concentrations of Pex5-cargo. Our findings lead us to suggest a model in which LLPS of Pex5-cargo with Pex13 and Pex14 results in transient protein transport channels.
Topics: Intracellular Membranes; Membrane Proteins; Peroxins; Peroxisome-Targeting Signal 1 Receptor; Peroxisomes; Phase Transition; Protein Binding; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Intrinsically Disordered Proteins
PubMed: 37165185
DOI: 10.1038/s41586-023-06044-1 -
Structural basis of ER-associated protein degradation mediated by the Hrd1 ubiquitin ligase complex.Science (New York, N.Y.) Apr 2020Misfolded luminal endoplasmic reticulum (ER) proteins undergo ER-associated degradation (ERAD-L): They are retrotranslocated into the cytosol, polyubiquitinated, and...
Misfolded luminal endoplasmic reticulum (ER) proteins undergo ER-associated degradation (ERAD-L): They are retrotranslocated into the cytosol, polyubiquitinated, and degraded by the proteasome. ERAD-L is mediated by the Hrd1 complex (composed of Hrd1, Hrd3, Der1, Usa1, and Yos9), but the mechanism of retrotranslocation remains mysterious. Here, we report a structure of the active Hrd1 complex, as determined by cryo-electron microscopy analysis of two subcomplexes. Hrd3 and Yos9 jointly create a luminal binding site that recognizes glycosylated substrates. Hrd1 and the rhomboid-like Der1 protein form two "half-channels" with cytosolic and luminal cavities, respectively, and lateral gates facing one another in a thinned membrane region. These structures, along with crosslinking and molecular dynamics simulation results, suggest how a polypeptide loop of an ERAD-L substrate moves through the ER membrane.
Topics: Carrier Proteins; Cryoelectron Microscopy; Endoplasmic Reticulum; Endoplasmic Reticulum-Associated Degradation; Membrane Glycoproteins; Membrane Proteins; Molecular Dynamics Simulation; Multiprotein Complexes; Protein Domains; Protein Folding; Proteolysis; Saccharomyces cerevisiae Proteins; Ubiquitin-Protein Ligases
PubMed: 32327568
DOI: 10.1126/science.aaz2449 -
Nature Reviews. Molecular Cell Biology Jun 2024Over the past two decades, protein S-acylation (often referred to as S-palmitoylation) has emerged as an important regulator of vital signalling pathways. S-Acylation is... (Review)
Review
Over the past two decades, protein S-acylation (often referred to as S-palmitoylation) has emerged as an important regulator of vital signalling pathways. S-Acylation is a reversible post-translational modification that involves the attachment of a fatty acid to a protein. Maintenance of the equilibrium between protein S-acylation and deacylation has demonstrated profound effects on various cellular processes, including innate immunity, inflammation, glucose metabolism and fat metabolism, as well as on brain and heart function. This Review provides an overview of current understanding of S-acylation and deacylation enzymes, their spatiotemporal regulation by sophisticated multilayered mechanisms, and their influence on protein function, cellular processes and physiological pathways. Furthermore, we examine how disruptions in protein S-acylation are associated with a broad spectrum of diseases from cancer to autoinflammatory disorders and neurological conditions.
Topics: Humans; Animals; Acylation; Protein Processing, Post-Translational; Signal Transduction; Lipoylation; Proteins
PubMed: 38355760
DOI: 10.1038/s41580-024-00700-8 -
Molecular Cell Aug 2020Transcription factors (TFs) that bind common DNA motifs in vitro occupy distinct sets of promoters in vivo, raising the question of how binding specificity is...
Transcription factors (TFs) that bind common DNA motifs in vitro occupy distinct sets of promoters in vivo, raising the question of how binding specificity is achieved. TFs are enriched with intrinsically disordered regions (IDRs). Such regions commonly form promiscuous interactions, yet their unique properties might also benefit specific binding-site selection. We examine this using Msn2 and Yap1, TFs of distinct families that contain long IDRs outside their DNA-binding domains. We find that these IDRs are both necessary and sufficient for localizing to the majority of target promoters. This IDR-directed binding does not depend on any localized domain but results from a multitude of weak determinants distributed throughout the entire IDR sequence. Furthermore, IDR specificity is conserved between distant orthologs, suggesting direct interaction with multiple promoters. We propose that distribution of sensing determinants along extended IDRs accelerates binding-site detection by rapidly localizing TFs to broad DNA regions surrounding these sites.
Topics: Binding Sites; Computational Biology; Conserved Sequence; DNA-Binding Proteins; Gene Expression Regulation, Fungal; Intrinsically Disordered Proteins; Models, Statistical; Nucleotide Motifs; Promoter Regions, Genetic; Protein Binding; Protein Interaction Domains and Motifs; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Sequence Deletion; Signal Transduction; Transcription Factors
PubMed: 32553192
DOI: 10.1016/j.molcel.2020.05.032 -
Biomolecules Apr 2023In this review, we present a comprehensive list of the ubiquitin-like modifiers (Ubls) of , a common model organism used to study fundamental cellular processes that are... (Review)
Review
In this review, we present a comprehensive list of the ubiquitin-like modifiers (Ubls) of , a common model organism used to study fundamental cellular processes that are conserved in complex multicellular organisms, such as humans. Ubls are a family of proteins that share structural relationships with ubiquitin, and which modify target proteins and lipids. These modifiers are processed, activated and conjugated to substrates by cognate enzymatic cascades. The attachment of substrates to Ubls alters the various properties of these substrates, such as function, interaction with the environment or turnover, and accordingly regulate key cellular processes, including DNA damage, cell cycle progression, metabolism, stress response, cellular differentiation, and protein homeostasis. Thus, it is not surprising that Ubls serve as tools to study the underlying mechanism involved in cellular health. We summarize current knowledge on the activity and mechanism of action of the Rub1, Smt3, Atg8, Atg12, Urm1 and Hub1 modifiers, all of which are highly conserved in organisms from yeast to humans.
Topics: Humans; Ubiquitins; Saccharomyces cerevisiae; Ubiquitin; Proteins; DNA Damage; Saccharomyces cerevisiae Proteins; Ligases
PubMed: 37238603
DOI: 10.3390/biom13050734 -
Biochimica Et Biophysica Acta. Gene... Feb 2021Histone post-translational modifications are essential for the regulation of gene expression in eukaryotes. Gcn5 (KAT2A) is a histone acetyltransferase that catalyzes... (Review)
Review
Histone post-translational modifications are essential for the regulation of gene expression in eukaryotes. Gcn5 (KAT2A) is a histone acetyltransferase that catalyzes the post-translational modification at multiple positions of histone H3 through the transfer of acetyl groups to the free amino group of lysine residues. Gcn5 catalyzes histone acetylation in the context of a HAT module containing the Ada2, Ada3 and Sgf29 subunits of the parent megadalton SAGA transcriptional coactivator complex. Biochemical and structural studies have elucidated mechanisms for Gcn5's acetyl- and other acyltransferase activities on histone substrates, for histone H3 phosphorylation and histone H3 methylation crosstalks with histone H3 acetylation, and for how Ada2 increases Gcn5's histone acetyltransferase activity. Other studies have identified Ada2 isoforms in SAGA-related complexes and characterized variant Gcn5 HAT modules containing these Ada2 isoforms. In this review, we highlight biochemical and structural studies of Gcn5 and its functional interactions with Ada2, Ada3 and Sgf29.
Topics: Acetylation; Cryoelectron Microscopy; Histone Acetyltransferases; Histones; Isoenzymes; Methylation; Multienzyme Complexes; Phosphorylation; Protein Processing, Post-Translational; Saccharomyces cerevisiae Proteins; Transcription Factors; p300-CBP Transcription Factors
PubMed: 32890768
DOI: 10.1016/j.bbagrm.2020.194629 -
Current Opinion in Structural Biology Feb 2020Proteins are subject to various conflicting forces that trade-off against each other. For example, during folding, the protein achieves lower enthalpy at the cost of... (Review)
Review
Proteins are subject to various conflicting forces that trade-off against each other. For example, during folding, the protein achieves lower enthalpy at the cost of lower entropy. Similarly, the trade-off for increased stability may be decreased flexibility, which may abolish allosteric pathways. Accordingly, stability trades-off against function, which may also trade-off against folding kinetics and mechanism. Furthermore, attaining increased stability may reduce a protein's ability to adopt novel functions. Understanding the biophysics and function of proteins requires quantification of the driving forces involved in each of the trade-offs. Indeed, quantification of the linkages in the network of trade-offs is essential to obtaining a more complete understanding of protein structure and function.
Topics: Animals; Entropy; Humans; Kinetics; Protein Folding; Proteins
PubMed: 31816559
DOI: 10.1016/j.sbi.2019.11.005 -
Protein and Peptide Letters 2023Proteins are essential biomacromolecules in all living systems because they are the prominent ultimate executives of the genetic information stored in DNA. Thus,... (Review)
Review
Proteins are essential biomacromolecules in all living systems because they are the prominent ultimate executives of the genetic information stored in DNA. Thus, studying protein is one of the central tasks in biological sciences. The complexity, diversity, and dynamics of a protein's structure, function, and structure-function relationship, the inherent structural fragility and thus the requirements on handling proteins to maintain protein's structural and functional orderliness make it a rather tricky task to work with protein. The approach to understanding the functions of a protein has been progressing steadily. In this paper, we reviewed the progress on the approach to the functional study of proteins that tremendously contributed to understanding their biological significance. Emphasis was put on the advances in the age in which high-throughput DNA sequencing and bioinformatics analysis are revolutionizing biological study.
Topics: Proteins; Biology
PubMed: 37151077
DOI: 10.2174/0929866530666230507212638 -
Advances in Biological Regulation May 2022The PAH1-encoded phosphatidate phosphatase, which catalyzes the dephosphorylation of phosphatidate to produce diacylglycerol, controls the divergence of phosphatidate... (Review)
Review
The PAH1-encoded phosphatidate phosphatase, which catalyzes the dephosphorylation of phosphatidate to produce diacylglycerol, controls the divergence of phosphatidate into triacylglycerol synthesis and phospholipid synthesis. Pah1 is inactive in the cytosol as a phosphorylated form and becomes active on the nuclear/endoplasmic reticulum membrane as a dephosphorylated form by the Nem1-Spo7 protein phosphatase complex. The phosphorylation of Pah1 by protein kinases, which include casein kinases I and II, Pho85-Pho80, Cdc28-cyclin B, and protein kinases A and C, controls its cellular location, catalytic activity, and susceptibility to proteasomal degradation. Nem1 (catalytic subunit) and Spo7 (regulatory subunit), which form a protein phosphatase complex catalyzing the dephosphorylation of Pah1 for its activation, are phosphorylated by protein kinases A and C. In this review, we discuss the functions and interrelationships of the protein kinases in the control of the Nem1-Spo7/Pah1 phosphatase cascade and lipid synthesis.
Topics: Lipids; Membrane Proteins; Nuclear Proteins; Phosphatidate Phosphatase; Phosphorylation; Protein Kinases; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 35231723
DOI: 10.1016/j.jbior.2022.100889