-
International Journal of Molecular... Jul 2020A delicate intracellular balance among protein synthesis, folding, and degradation is essential to maintaining protein homeostasis or proteostasis, and it is challenged... (Review)
Review
A delicate intracellular balance among protein synthesis, folding, and degradation is essential to maintaining protein homeostasis or proteostasis, and it is challenged by genetic and environmental factors. Molecular chaperones and the ubiquitin proteasome system (UPS) play a vital role in proteostasis for normal cellular function. As part of protein quality control, molecular chaperones recognize misfolded proteins and assist in their refolding. Proteins that are beyond repair or refolding undergo degradation, which is largely mediated by the UPS. The importance of protein quality control is becoming ever clearer, but it can also be a disease-causing mechanism. Diseases such as phenylketonuria (PKU) and hereditary tyrosinemia-I (HT1) are caused due to mutations in and gene, resulting in reduced protein stability, misfolding, accelerated degradation, and deficiency in functional proteins. Misfolded or partially unfolded proteins do not necessarily lose their functional activity completely. Thus, partially functional proteins can be rescued from degradation by molecular chaperones and deubiquitinating enzymes (DUBs). Deubiquitination is an important mechanism of the UPS that can reverse the degradation of a substrate protein by covalently removing its attached ubiquitin molecule. In this review, we discuss the importance of molecular chaperones and DUBs in reducing the severity of PKU and HT1 by stabilizing and rescuing mutant proteins.
Topics: Animals; Deubiquitinating Enzymes; Humans; Molecular Chaperones; Phenylketonurias; Proteasome Endopeptidase Complex; Protein Folding; Protein Stability; Proteolysis; Tyrosinemias; Ubiquitination
PubMed: 32679806
DOI: 10.3390/ijms21144996 -
International Journal of Molecular... Feb 2021Aggrephagy is defined as the selective degradation of aggregated proteins by autophagosomes. Protein aggregation in organs and cells has been highlighted as a cause of... (Review)
Review
Aggrephagy is defined as the selective degradation of aggregated proteins by autophagosomes. Protein aggregation in organs and cells has been highlighted as a cause of multiple diseases, including neurodegenerative diseases, cardiac failure, and renal failure. Aggregates could pose a hazard for cell survival. Cells exhibit three main mechanisms against the accumulation of aggregates: protein refolding by upregulation of chaperones, reduction of protein overload by translational inhibition, and protein degradation by the ubiquitin-proteasome and autophagy-lysosome systems. Deletion of autophagy-related genes reportedly contributes to intracellular protein aggregation in vivo. Some proteins recognized in aggregates in preeclamptic placentas include those involved in neurodegenerative diseases. As aggregates are derived both intracellularly and extracellularly, special endocytosis for extracellular aggregates also employs the autophagy machinery. In this review, we discuss how the deficiency of aggrephagy and/or macroautophagy leads to poor placentation, resulting in preeclampsia or fetal growth restriction.
Topics: Animals; Female; Humans; Lysosomes; Macroautophagy; Placenta; Pre-Eclampsia; Pregnancy; Protein Aggregation, Pathological
PubMed: 33670947
DOI: 10.3390/ijms22052432 -
Journal of Colloid and Interface Science May 2024Nonionic surfactants can counter the deleterious effect that anionic surfactants have on proteins, where the folded states are retrieved from a previously unfolded...
HYPOTHESIS
Nonionic surfactants can counter the deleterious effect that anionic surfactants have on proteins, where the folded states are retrieved from a previously unfolded state. However, further studies are required to refine our understanding of the underlying mechanism of the refolding process. While interactions between nonionic surfactants and tightly folded proteins are not anticipated, we hypothesized that intermediate stages of surfactant-induced unfolding could define new interaction mechanisms by which nonionic surfactants can further alter protein conformation.
EXPERIMENTS
In this work, the behavior of three model proteins (human growth hormone, bovine serum albumin, and β-lactoglobulin) was investigated in the presence of the anionic surfactant sodium dodecylsulfate, the nonionic surfactant β-dodecylmaltoside, and mixtures of both surfactants. The transitions occurring to the proteins were determined using intrinsic fluorescence spectroscopy and far-UV circular dichroism. Based on these results, we developed a detailed interaction model for human growth hormone. Using nuclear magnetic resonance and contrast-variation small-angle neutron scattering, we studied the amino acid environment and the conformational state of the protein.
FINDINGS
The results demonstrate the key role of surfactant cooperation in defining the conformational state of the proteins, which can shift away or toward the folded state depending on the nonionic-to-ionic surfactant ratio. Dodecylmaltoside, initially a non-interacting surfactant, can unexpectedly associate with sodium dodecylsulfate-unfolded proteins to further impact their conformation at low nonionic-to-ionic surfactant ratio. When this ratio increases, the protein begins to retrieve the folded state. However, the native conformation cannot be fully recovered due to remnant surfactant molecules still adsorbed to the protein. This study demonstrates that the conformational landscape of the protein depends on a delicate interplay between the surfactants, ultimately controlled by the ratio between them, resulting in unpredictable changes in the protein conformation.
PubMed: 38838632
DOI: 10.1016/j.jcis.2024.05.157 -
Molecules (Basel, Switzerland) Mar 2023Serratiopeptidase is a clinical therapeutic protein for the treatment of human diseases such as arthritis, bronchitis, and thrombosis. Yet production of this protein in...
Serratiopeptidase is a clinical therapeutic protein for the treatment of human diseases such as arthritis, bronchitis, and thrombosis. Yet production of this protein in a heterologous host (e.g., ) is difficult due to the issue of protein insolubility and the requirement of laborious refolding procedures. Cell-free protein synthesis (CFPS) systems, derived from crude cell extracts, are effective platforms for the expression of recombinant proteins in vitro. Here, we report a new method to produce serratiopeptidase by using an -based CFPS system. After rational selection of cell extracts and construction of expression vectors, soluble expression of serratiopeptidase was achieved and the enzyme activity could be readily tested in the cell-free reaction mixture. By further optimizing the key parameters, optimum conditions for the enzyme activity assay were obtained, including the pH value at 5, reaction temperature at 45 °C, substrate concentration at 10 mg/mL, and supplementing Ca ions at 5 mM. Moreover, the CFPS mixture was freeze-dried and the activity of serratiopeptidase could be regenerated by hydration without losing activity. Overall, the CFPS system enabled soluble expression of serratiopeptidase with catalytic activity, providing a new and promising approach for this enzyme production. Our work extends the utility of the cell-free platform to produce therapeutic proteins with clinical applications.
Topics: Humans; Escherichia coli; Cell Extracts; Protein Biosynthesis; Recombinant Proteins; Cell-Free System
PubMed: 37049893
DOI: 10.3390/molecules28073132 -
International Journal of Molecular... Jan 2022The prevailing current view of protein folding is the thermodynamic hypothesis, under which the native folded conformation of a protein corresponds to the global minimum... (Review)
Review
The prevailing current view of protein folding is the thermodynamic hypothesis, under which the native folded conformation of a protein corresponds to the global minimum of Gibbs free energy . We question this concept and show that the empirical evidence behind the thermodynamic hypothesis of folding is far from strong. Furthermore, physical theory-based approaches to the prediction of protein folds and their folding pathways so far have invariably failed except for some very small proteins, despite decades of intensive theory development and the enormous increase of computer power. The recent spectacular successes in protein structure prediction owe to evolutionary modeling of amino acid sequence substitutions enhanced by deep learning methods, but even these breakthroughs provide no information on the protein folding mechanisms and pathways. We discuss an alternative view of protein folding, under which the native state of most proteins does not occupy the global free energy minimum, but rather, a local minimum on a fluctuating free energy landscape. We further argue that Δ of folding is likely to be positive for the majority of proteins, which therefore fold into their native conformations only through interactions with the energy-dependent molecular machinery of living cells, in particular, the translation system and chaperones. Accordingly, protein folding should be modeled as it occurs in vivo, that is, as a non-equilibrium, active, energy-dependent process.
Topics: Algorithms; Kinetics; Models, Molecular; Models, Theoretical; Protein Conformation; Protein Folding; Protein Refolding; Protein Stability; Proteins; Proteome; Proteomics; Recombinant Proteins; Solubility; Species Specificity; Thermodynamics
PubMed: 35008947
DOI: 10.3390/ijms23010521 -
Journal of Parasitic Diseases :... Dec 2020has the most adenine (A)- and thymine (T)-rich genome known to date, and 24-30% of the proteome contains asparagine (N) and glutamine (Q) residues. In general,... (Review)
Review
has the most adenine (A)- and thymine (T)-rich genome known to date, and 24-30% of the proteome contains asparagine (N) and glutamine (Q) residues. In general, asparagine repeats in proteins increase the propensity for aggregation, especially at elevated temperatures, which occur routinely in parasites during exoerythrocytic and erythrocytic developmental stages in a vertebrate host. The exported chaperone machinery is comprised of an exported PfHsp70-x protein and its co-chaperone PfHsp40-x1 in the host erythrocyte compartment, and PfHsp70-z and its co-chaperones in the parasite cytoplasm have been identified. In vitro assays reveal that these chaperone partners function in refolding of asparagine-rich polypeptides. The identification and chaperoning of exported poly-asparagine-containing proteins, and the biological roles and the protection mechanisms of during febrile conditions by the exported chaperone machinery are discussed.
PubMed: 33184535
DOI: 10.1007/s12639-020-01251-3 -
The FEBS Journal Mar 2021Unfolding and refolding of multidomain proteins under force have yet to be recognized as a major mechanism of function for proteins in vivo. In this review, we discuss... (Review)
Review
Unfolding and refolding of multidomain proteins under force have yet to be recognized as a major mechanism of function for proteins in vivo. In this review, we discuss the inherent properties of multidomain proteins under a force vector from a structural and functional perspective. We then characterize three main systems where multidomain proteins could play major roles through mechanical unfolding: muscular contraction, cellular mechanotransduction, and bacterial adhesion. We analyze how key multidomain proteins for each system can produce a gain-of-function from the perspective of a fine-tuned quantized response, a molecular battery, delivery of mechanical work through refolding, elasticity tuning, protection and exposure of cryptic sites, and binding-induced mechanical changes. Understanding how mechanical unfolding and refolding affect function will have important implications in designing mechano-active drugs against conditions such as muscular dystrophy, cancer, or novel antibiotics.
Topics: Elasticity; Mechanotransduction, Cellular; Models, Molecular; Protein Binding; Protein Domains; Protein Folding; Protein Unfolding; Proteins; Stress, Mechanical; Thermodynamics
PubMed: 32761965
DOI: 10.1111/febs.15508 -
Frontiers in Molecular Biosciences 2023While proteins populating their native conformations constitute the functional entities of cells, protein aggregates are traditionally associated with cellular... (Review)
Review
While proteins populating their native conformations constitute the functional entities of cells, protein aggregates are traditionally associated with cellular dysfunction, stress and disease. During recent years, it has become clear that large aggregate-like protein condensates formed liquid-liquid phase separation age into more solid aggregate-like particles that harbor misfolded proteins and are decorated by protein quality control factors. The constituent proteins of the condensates/aggregates are disentangled by protein disaggregation systems mainly based on Hsp70 and AAA ATPase Hsp100 chaperones prior to their handover to refolding and degradation systems. Here, we discuss the functional roles that condensate formation/aggregation and disaggregation play in protein quality control to maintain proteostasis and why it matters for understanding health and disease.
PubMed: 37021114
DOI: 10.3389/fmolb.2023.1155521 -
Frontiers in Aging 2022Cardiovascular disorder is the major health burden and cause of death among individuals worldwide. As the cardiomyocytes lack the ability for self-renewal, it is utmost... (Review)
Review
Cardiovascular disorder is the major health burden and cause of death among individuals worldwide. As the cardiomyocytes lack the ability for self-renewal, it is utmost necessary to surveil the protein quality in the cells. The Bcl-2 associated anthanogene protein (BAG) family and molecular chaperones (HSP70, HSP90) actively participate in maintaining cellular protein quality control (PQC) to limit cellular dysfunction in the cells. The BAG family contains a unique BAG domain which facilitates their interaction with the ATPase domain of the heat shock protein 70 (HSP70) to assist in protein folding. Among the BAG family members (BAG1-6), BAG5 protein is unique since it has five domains in tandem, and the binding of BD5 induces certain conformational changes in the nucleotide-binding domain (NBD) of HSP70 such that it loses its affinity for binding to ADP and results in enhanced protein refolding activity of HSP70. In this review, we shall describe the role of BAG5 in modulating mitophagy, endoplasmic stress, and cellular viability. Also, we have highlighted the interaction of BAG5 with other proteins, including PINK, DJ-1, CHIP, and their role in cellular PQC. Apart from this, we have described the role of BAG5 in cellular metabolism and aging.
PubMed: 35821856
DOI: 10.3389/fragi.2022.844168 -
Cellular and Molecular Life Sciences :... Dec 2021Accumulation of misfolded proteins in the endoplasmic reticulum (ER) induces a well-orchestrated cellular response to reduce the protein burden within the ER. This... (Review)
Review
Accumulation of misfolded proteins in the endoplasmic reticulum (ER) induces a well-orchestrated cellular response to reduce the protein burden within the ER. This unfolded protein response (UPR) is controlled primarily by three transmembrane proteins, IRE1α, ATF6, and PERK, the activity of which is controlled by BiP, the ER-resident Hsp70 protein. Binding of BiP to co-chaperones via their highly conserved J-domains stimulates the intrinsic ATPase activity of BiP, thereby providing the energy necessary for (re-)folding of proteins, or for targeting of misfolded proteins to the degradation pathway, processes specified and controlled by the respective co-chaperone. In this review, our aim is to elucidate the function of the co-chaperone ERDJ4, also known as MDG1, MDJ7, or DNAJB9. Knockout and knockin experiments clearly point to the central role of ERDJ4 in controlling lipogenesis and protein synthesis by promoting degradation of SREBP1c and the assembly of the protein complex mTORC2. Accumulating data reveal that ERDJ4 controls epithelial-to-mesenchymal transition, a central process during embryogenesis, in wound healing, and tumor development. Overexpression of ERdj4 has been shown to improve engraftment of transplanted human stem cells, possibly due to its ability to promote cellular survival in stressed cells. High ERDJ4-plasma levels are specific for fibrillary glomerulonephritis and serve as a diagnostic marker. As outlined in this review, the functions of ERDJ4 are manifold, depending on the cellular (patho-) physiological state, the cellular protein repertoire, and the subcellular localization of ERDJ4.
Topics: Animals; Apoptosis; Biomarkers; Endoplasmic Reticulum Stress; Epithelial-Mesenchymal Transition; Humans; Molecular Chaperones; Protein Folding
PubMed: 34950970
DOI: 10.1007/s00018-021-04082-4