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Nature Reviews. Molecular Cell Biology Dec 2023Despite advances in machine learning-based protein structure prediction, we are still far from fully understanding how proteins fold into their native conformation. The... (Review)
Review
Despite advances in machine learning-based protein structure prediction, we are still far from fully understanding how proteins fold into their native conformation. The conventional notion that polypeptides fold spontaneously to their biologically active states has gradually been replaced by our understanding that cellular protein folding often requires context-dependent guidance from molecular chaperones in order to avoid misfolding. Misfolded proteins can aggregate into larger structures, such as amyloid fibrils, which perpetuate the misfolding process, creating a self-reinforcing cascade. A surge in amyloid fibril structures has deepened our comprehension of how a single polypeptide sequence can exhibit multiple amyloid conformations, known as polymorphism. The assembly of these polymorphs is not a random process but is influenced by the specific conditions and tissues in which they originate. This observation suggests that, similar to the folding of native proteins, the kinetics of pathological amyloid assembly are modulated by interactions specific to cells and tissues. Here, we review the current understanding of how intrinsic protein conformational propensities are modulated by physiological and pathological interactions in the cell to shape protein misfolding and aggregation pathology.
Topics: Protein Conformation; Protein Folding; Amyloid; Peptides; Molecular Chaperones
PubMed: 37684425
DOI: 10.1038/s41580-023-00647-2 -
Nature Sep 2023Abnormal assembly of tau, α-synuclein, TDP-43 and amyloid-β proteins into amyloid filaments defines most human neurodegenerative diseases. Genetics provides a direct... (Review)
Review
Abnormal assembly of tau, α-synuclein, TDP-43 and amyloid-β proteins into amyloid filaments defines most human neurodegenerative diseases. Genetics provides a direct link between filament formation and the causes of disease. Developments in cryo-electron microscopy (cryo-EM) have made it possible to determine the atomic structures of amyloids from postmortem human brains. Here we review the structures of brain-derived amyloid filaments that have been determined so far and discuss their impact on research into neurodegeneration. Whereas a given protein can adopt many different filament structures, specific amyloid folds define distinct diseases. Amyloid structures thus provide a description of neuropathology at the atomic level and a basis for studying disease. Future research should focus on model systems that replicate the structures observed in disease to better understand the molecular mechanisms of disease and develop improved diagnostics and therapies.
Topics: Humans; alpha-Synuclein; Amyloid; Amyloid beta-Peptides; Cryoelectron Microscopy; Neurodegenerative Diseases; Pathology, Molecular; Protein Folding
PubMed: 37758888
DOI: 10.1038/s41586-023-06437-2 -
Nature Aug 2023Advances in DNA sequencing and machine learning are providing insights into protein sequences and structures on an enormous scale. However, the energetics driving...
Advances in DNA sequencing and machine learning are providing insights into protein sequences and structures on an enormous scale. However, the energetics driving folding are invisible in these structures and remain largely unknown. The hidden thermodynamics of folding can drive disease, shape protein evolution and guide protein engineering, and new approaches are needed to reveal these thermodynamics for every sequence and structure. Here we present cDNA display proteolysis, a method for measuring thermodynamic folding stability for up to 900,000 protein domains in a one-week experiment. From 1.8 million measurements in total, we curated a set of around 776,000 high-quality folding stabilities covering all single amino acid variants and selected double mutants of 331 natural and 148 de novo designed protein domains 40-72 amino acids in length. Using this extensive dataset, we quantified (1) environmental factors influencing amino acid fitness, (2) thermodynamic couplings (including unexpected interactions) between protein sites, and (3) the global divergence between evolutionary amino acid usage and protein folding stability. We also examined how our approach could identify stability determinants in designed proteins and evaluate design methods. The cDNA display proteolysis method is fast, accurate and uniquely scalable, and promises to reveal the quantitative rules for how amino acid sequences encode folding stability.
Topics: Amino Acids; Biology; DNA, Complementary; Protein Folding; Protein Stability; Proteins; Thermodynamics; Proteolysis; Protein Engineering; Protein Domains; Mutation
PubMed: 37468638
DOI: 10.1038/s41586-023-06328-6 -
Current Opinion in Structural Biology Jun 2024
Topics: Protein Folding; Protein Binding; Proteins; Humans
PubMed: 38417184
DOI: 10.1016/j.sbi.2024.102791 -
Nature Reviews. Molecular Cell Biology Nov 2023Heat shock protein 90 (HSP90) is a chaperone with vital roles in regulating proteostasis, long recognized for its function in protein folding and maturation. A view is... (Review)
Review
Heat shock protein 90 (HSP90) is a chaperone with vital roles in regulating proteostasis, long recognized for its function in protein folding and maturation. A view is emerging that identifies HSP90 not as one protein that is structurally and functionally homogeneous but, rather, as a protein that is shaped by its environment. In this Review, we discuss evidence of multiple structural forms of HSP90 in health and disease, including homo-oligomers and hetero-oligomers, also termed epichaperomes, and examine the impact of stress, post-translational modifications and co-chaperones on their formation. We describe how these variations influence context-dependent functions of HSP90 as well as its interaction with other chaperones, co-chaperones and proteins, and how this structural complexity of HSP90 impacts and is impacted by its interaction with small molecule modulators. We close by discussing recent developments regarding the use of HSP90 inhibitors in cancer and how our new appreciation of the structural and functional heterogeneity of HSP90 invites a re-evaluation of how we discover and implement HSP90 therapeutics for disease treatment.
Topics: HSP90 Heat-Shock Proteins; Molecular Chaperones; Protein Folding; Proteostasis; Homeostasis
PubMed: 37524848
DOI: 10.1038/s41580-023-00640-9 -
Nature Aug 2023Dominant optic atrophy is one of the leading causes of childhood blindness. Around 60-80% of cases are caused by mutations of the gene that encodes optic atrophy protein...
Dominant optic atrophy is one of the leading causes of childhood blindness. Around 60-80% of cases are caused by mutations of the gene that encodes optic atrophy protein 1 (OPA1), a protein that has a key role in inner mitochondrial membrane fusion and remodelling of cristae and is crucial for the dynamic organization and regulation of mitochondria. Mutations in OPA1 result in the dysregulation of the GTPase-mediated fusion process of the mitochondrial inner and outer membranes. Here we used cryo-electron microscopy methods to solve helical structures of OPA1 assembled on lipid membrane tubes, in the presence and absence of nucleotide. These helical assemblies organize into densely packed protein rungs with minimal inter-rung connectivity, and exhibit nucleotide-dependent dimerization of the GTPase domains-a hallmark of the dynamin superfamily of proteins. OPA1 also contains several unique secondary structures in the paddle domain that strengthen its membrane association, including membrane-inserting helices. The structural features identified in this study shed light on the effects of pathogenic point mutations on protein folding, inter-protein assembly and membrane interactions. Furthermore, mutations that disrupt the assembly interfaces and membrane binding of OPA1 cause mitochondrial fragmentation in cell-based assays, providing evidence of the biological relevance of these interactions.
Topics: Cryoelectron Microscopy; GTP Phosphohydrolases; Membrane Fusion; Mitochondria; Mitochondrial Dynamics; Mitochondrial Membranes; Mutation; Nucleotides; Protein Binding; Protein Domains; Protein Folding; Protein Multimerization; Protein Structure, Secondary; Humans
PubMed: 37612506
DOI: 10.1038/s41586-023-06462-1 -
Journal of Molecular Biology Jul 2024The mRNA coding sequence defines not only the amino acid sequence of the protein, but also the speed at which the ribosomes move along the mRNA while making the protein.... (Review)
Review
The mRNA coding sequence defines not only the amino acid sequence of the protein, but also the speed at which the ribosomes move along the mRNA while making the protein. The non-uniform local kinetics - denoted as translational rhythm - is similar among mRNAs coding for related protein folds. Deviations from this conserved rhythm can result in protein misfolding. In this review we summarize the experimental evidence demonstrating how local translation rates affect cotranslational protein folding, with the focus on the synonymous codons and patches of charged residues in the nascent peptide as best-studied examples. Alterations in nascent protein conformations due to disturbed translational rhythm can persist off the ribosome, as demonstrated by the effects of synonymous codon variants of several disease-related proteins. Charged amino acid patches in nascent chains also modulate translation and cotranslational protein folding, and can abrogate translation when placed at the N-terminus of the nascent peptide. During cotranslational folding, incomplete nascent chains navigate through a unique conformational landscape in which earlier intermediate states become inaccessible as the nascent peptide grows. Precisely tuned local translation rates, as well as interactions with the ribosome, guide the folding pathway towards the native structure, whereas deviations from the natural translation rhythm may favor pathways leading to trapped misfolded states. Deciphering the 'folding code' of the mRNA will contribute to understanding the diseases caused by protein misfolding and to rational protein design.
Topics: Protein Folding; Protein Biosynthesis; Ribosomes; Humans; RNA, Messenger; Proteins; Codon; Protein Conformation; Kinetics; Animals
PubMed: 38065274
DOI: 10.1016/j.jmb.2023.168384 -
Molecular Cell Dec 2023N-glycans act as quality control tags by recruiting lectin chaperones to assist protein maturation in the endoplasmic reticulum. The location and composition of...
N-glycans act as quality control tags by recruiting lectin chaperones to assist protein maturation in the endoplasmic reticulum. The location and composition of N-glycans (glyco-code) are key to the chaperone-selection process. Serpins, a class of serine protease inhibitors, fold non-sequentially to achieve metastable active states. Here, the role of the glyco-code in assuring successful maturation and quality control of two human serpins, alpha-1 antitrypsin (AAT) and antithrombin III (ATIII), is described. We find that AAT, which has glycans near its N terminus, is assisted by early lectin chaperone binding. In contrast, ATIII, which has more C-terminal glycans, is initially helped by BiP and then later by lectin chaperones mediated by UGGT reglucosylation. UGGT action is increased for misfolding-prone disease variants, and these clients are preferentially glucosylated on their most C-terminal glycan. Our study illustrates how serpins utilize N-glycan presence, position, and composition to direct their proper folding, quality control, and trafficking.
Topics: Humans; Molecular Chaperones; Protein Folding; Lectins; Polysaccharides; Quality Control
PubMed: 38052210
DOI: 10.1016/j.molcel.2023.11.006 -
Nature Jan 2024AlphaFold2 (ref. ) has revolutionized structural biology by accurately predicting single structures of proteins. However, a protein's biological function often depends...
AlphaFold2 (ref. ) has revolutionized structural biology by accurately predicting single structures of proteins. However, a protein's biological function often depends on multiple conformational substates, and disease-causing point mutations often cause population changes within these substates. We demonstrate that clustering a multiple-sequence alignment by sequence similarity enables AlphaFold2 to sample alternative states of known metamorphic proteins with high confidence. Using this method, named AF-Cluster, we investigated the evolutionary distribution of predicted structures for the metamorphic protein KaiB and found that predictions of both conformations were distributed in clusters across the KaiB family. We used nuclear magnetic resonance spectroscopy to confirm an AF-Cluster prediction: a cyanobacteria KaiB variant is stabilized in the opposite state compared with the more widely studied variant. To test AF-Cluster's sensitivity to point mutations, we designed and experimentally verified a set of three mutations predicted to flip KaiB from Rhodobacter sphaeroides from the ground to the fold-switched state. Finally, screening for alternative states in protein families without known fold switching identified a putative alternative state for the oxidoreductase Mpt53 in Mycobacterium tuberculosis. Further development of such bioinformatic methods in tandem with experiments will probably have a considerable impact on predicting protein energy landscapes, essential for illuminating biological function.
Topics: Cluster Analysis; Mutation; Protein Conformation; Proteins; Sequence Alignment; Machine Learning; Rhodobacter sphaeroides; Bacterial Proteins; Protein Folding
PubMed: 37956700
DOI: 10.1038/s41586-023-06832-9 -
Current Opinion in Structural Biology Dec 2023Topologically knotted proteins have entangled structural elements within their native structures that cannot be disentangled simply by pulling from the N- and C-termini.... (Review)
Review
Topologically knotted proteins have entangled structural elements within their native structures that cannot be disentangled simply by pulling from the N- and C-termini. Systematic surveys have identified different types of knotted protein structures, constituting as much as 1% of the total entries within the Protein Data Bank. Many knotted proteins rely on their knotted structural elements to carry out evolutionarily conserved biological functions. Being knotted may also provide mechanical stability to withstand unfolding-coupled proteolysis. Reconfiguring a knotted protein topology by circular permutation or cyclization provides insights into the importance of being knotted in the context of folding and functions. With the explosion of predicted protein structures by artificial intelligence, we are now entering a new era of exploring the entangled protein universe.
Topics: Protein Folding; Artificial Intelligence; Proteins; Protein Conformation
PubMed: 37778185
DOI: 10.1016/j.sbi.2023.102709