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Dual Enkephalinase Inhibitors and Their Role in Chronic Pain Management.
Southerland, WA, Gillis, J, Kuppalli, S, Fonseca, A, Mendelson, A, Horine, SV, Bansal, N, Gulati, A
Current pain and headache reports. 2021;(5):29
Abstract
PURPOSE OF REVIEW Dual enkephalinase inhibitors (DENKIs) are pain medications that indirectly activate opioid receptors and can be used as an alternative to traditional opioids. Understanding the physiology of enkephalins and their inhibitors and the pharmacology of these drugs will allow for proper clinical application for chronic pain patients in the future. RECENT FINDINGS DENKIs can be used as an alternative mode of analgesia for patients suffering from chronic pain by preventing the degradation of endogenous opioid ligands. By inhibiting the two major enkephalin-degrading enzymes (neprilysin and aminopeptidase N), DENKIs can provide analgesia with less adverse effects than nonendogenous opioids. The purpose of this paper is to review the current literature investigating DENKIs and explore their contribution to chronic pain management.
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Harnessing the potential of bacterial oxidative folding to aid protein production.
Slater, SL, Mavridou, DAI
Molecular microbiology. 2021;(1):16-28
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Abstract
Protein folding is central to both biological function and recombinant protein production. In bacterial expression systems, which are easy to use and offer high protein yields, production of the protein of interest in its native fold can be hampered by the limitations of endogenous posttranslational modification systems. Disulfide bond formation, entailing the covalent linkage of proximal cysteine amino acids, is a fundamental posttranslational modification reaction that often underpins protein stability, especially in extracytoplasmic environments. When these bonds are not formed correctly, the yield and activity of the resultant protein are dramatically decreased. Although the mechanism of oxidative protein folding is well understood, unwanted or incorrect disulfide bond formation often presents a stumbling block for the expression of cysteine-containing proteins in bacteria. It is therefore important to consider the biochemistry of prokaryotic disulfide bond formation systems in the context of protein production, in order to take advantage of the full potential of such pathways in biotechnology applications. Here, we provide a critical overview of the use of bacterial oxidative folding in protein production so far, and propose a practical decision-making workflow for exploiting disulfide bond formation for the expression of any given protein of interest.
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Mechanisms of Disulfide Bond Formation in Nascent Polypeptides Entering the Secretory Pathway.
Robinson, PJ, Bulleid, NJ
Cells. 2020;(9)
Abstract
Disulphide bonds are an abundant feature of proteins across all domains of life that are important for structure, stability, and function. In eukaryotic cells, a major site of disulphide bond formation is the endoplasmic reticulum (ER). How cysteines correctly pair during polypeptide folding to form the native disulphide bond pattern is a complex problem that is not fully understood. In this paper, the evidence for different folding mechanisms involved in ER-localised disulphide bond formation is reviewed with emphasis on events that occur during ER entry. Disulphide formation in nascent polypeptides is discussed with focus on (i) its mechanistic relationship with conformational folding, (ii) evidence for its occurrence at the co-translational stage during ER entry, and (iii) the role of protein disulphide isomerase (PDI) family members. This review highlights the complex array of cellular processes that influence disulphide bond formation and identifies key questions that need to be addressed to further understand this fundamental process.
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Thiol-disulfide homeostasis: an integrated approach with biochemical and clinical aspects.
Erel, Ö, Erdoğan, S
Turkish journal of medical sciences. 2020;(SI-2):1728-1738
Abstract
Dynamic thiol-disulfide homeostasis (TDH) is a new area has begun to attract more scrutiny. Dynamic TDH is reversal of thiol oxidation in proteins and represents the status of thiols (-SH) and disulfides (-S-S-). Organic compounds containing the sulfhydryl group is called thiol, composed of sulfur and hydrogen atoms. Disulfides are the most important class of dynamic, redox responsive covalent bonds build in between two thiol groups. For many years, thiol levels were analyzed by several methods. During last years, measurements of disulfide levels have been analyzed by a novel automated method, developed by Erel and Neselioglu. In this method, addition to thiol (termed as native thiol) levels, disulfide levels were also measured and sum of native thiol and disulfide levels were termed as total thiol. Therefore, TDH was begun to be understood in organism. In healthy humans, TDH is maintained within a certain range. Dysregulated dynamic TDH has been implicated several disorders with unknown etiology. A growing body of evidence has demonstrated that the thiol-disulfide homeostasis is involved in variety diseases, such as diabetes mellitus, hypertension, nonsmall cell lung cancer, familial Mediterranean fever (FMF), inflammatory bowel diseases, occupational diseases, gestational diabetes mellitus and preeclampsia. These results may elucidate some pathogenic mechanism or may be a predictor indicating diagnostic clue, prognostic marker or therapeutic sign. In conclusion, protection of the thiol-disulfide homeostasis is of great importance for the human being. Evidence achieved so far has proposed that thiol-disulfide homeostasis is an important issue needs to elucidate wholly.
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Protein Disulfide Exchange by the Intramembrane Enzymes DsbB, DsbD, and CcdA.
Bushweller, JH
Journal of molecular biology. 2020;(18):5091-5103
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Abstract
The formation of disulfide bonds in proteins is an essential process in both prokaryotes and eukaryotes. In gram-negative bacteria including Escherichia coli, the proteins DsbA and DsbB mediate the formation of disulfide bonds in the periplasm. DsbA acts as the periplasmic oxidant of periplasmic substrate proteins. DsbA is reoxidized by transfer of reducing equivalents to the 4 TM helix membrane protein DsbB, which transfers reducing equivalents to ubiquinone or menaquinone. Multiple structural studies of DsbB have provided detailed structural information on intermediates in the process of DsbB catalyzed oxidation of DsbA. These structures and the insights gained are described. In proteins with more than one pair of Cys residues, there is the potential for formation of non-native disulfide bonds, making it necessary for the cell to have a mechanism for the isomerization of such non-native disulfide bonds. In E. coli, this is mediated by the proteins DsbC and DsbD. DsbC reduces mis-formed disulfide bonds. The eight-TM-helix protein DsbD reduces DsbC and is itself reduced by cytoplasmic thioredoxin. DsbD also contributes reducing equivalents for the reduction of cytochrome c to facilitate heme attachment. The DsbD functional homolog CcdA is a six-TM-helix membrane protein that provides reducing equivalents for the reduction of cytochrome c. A recent structure determination of CcdA has provided critical insights into how reducing equivalents are transferred across the membrane that likely also provides understanding how this is achieved by DsbD as well. This structure and the insights gained are described.
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Protein import by the mitochondrial disulfide relay in higher eukaryotes.
Finger, Y, Riemer, J
Biological chemistry. 2020;(6-7):749-763
Abstract
The proteome of the mitochondrial intermembrane space (IMS) contains more than 100 proteins, all of which are synthesized on cytosolic ribosomes and consequently need to be imported by dedicated machineries. The mitochondrial disulfide relay is the major import machinery for soluble proteins in the IMS. Its major component, the oxidoreductase MIA40, interacts with incoming substrates, retains them in the IMS, and oxidatively folds them. After this reaction, MIA40 is reoxidized by the sulfhydryl oxidase augmenter of liver regeneration, which couples disulfide formation by this machinery to the activity of the respiratory chain. In this review, we will discuss the import of IMS proteins with a focus on recent findings showing the diversity of disulfide relay substrates, describing the cytosolic control of this import system and highlighting the physiological relevance of the disulfide relay machinery in higher eukaryotes.
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Life inside and out: making and breaking protein disulfide bonds in Chlamydia.
Christensen, S, McMahon, RM, Martin, JL, Huston, WM
Critical reviews in microbiology. 2019;(1):33-50
Abstract
Disulphide bonds are widely used among all domains of life to provide structural stability to proteins and to regulate enzyme activity. Chlamydia spp. are obligate intracellular bacteria that are especially dependent on the formation and degradation of protein disulphide bonds. Members of the genus Chlamydia have a unique biphasic developmental cycle alternating between two distinct cell types; the extracellular infectious elementary body (EB) and the intracellular replicating reticulate body. The proteins in the envelope of the EB are heavily cross-linked with disulphides and this is known to be critical for this infectious phase. In this review, we provide a comprehensive summary of what is known about the redox state of chlamydial envelope proteins throughout the developmental cycle. We focus especially on the factors responsible for degradation and formation of disulphide bonds in Chlamydia and how this system compares with redox regulation in other organisms. Focussing on the unique biology of Chlamydia enables us to provide important insights into how specialized suites of disulphide bond (Dsb) proteins cater for specific bacterial environments and lifecycles.
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Applications of catalyzed cytoplasmic disulfide bond formation.
Saaranen, MJ, Ruddock, LW
Biochemical Society transactions. 2019;(5):1223-1231
Abstract
Disulfide bond formation is an essential post-translational modification required for many proteins to attain their native, functional structure. The formation of disulfide bonds, otherwise known as oxidative protein folding, occurs in the endoplasmic reticulum and mitochondrial inter-membrane space in eukaryotes and the periplasm of prokaryotes. While there are differences in the molecular mechanisms of oxidative folding in different compartments, it can essentially be broken down into two steps, disulfide formation and disulfide isomerization. For both steps, catalysts exist in all compartments where native disulfide bond formation occurs. Due to the importance of disulfide bonds for a plethora of proteins, considerable effort has been made to generate cell factories which can make them more efficiently and cheaper. Recently synthetic biology has been used to transfer catalysts of native disulfide bond formation into the cytoplasm of prokaryotes such as Escherichia coli. While these engineered systems cannot yet rival natural systems in the range and complexity of disulfide-bonded proteins that can be made, a growing range of proteins have been made successfully and yields of homogenously folded eukaryotic proteins exceeding g/l yields have been obtained. This review will briefly give an overview of such systems, the uses reported to date and areas of future potential development, including combining with engineered systems for cytoplasmic glycosylation.
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Engineering of the Dsb (disulfide bond) proteins - contribution towards understanding their mechanism of action and their applications in biotechnology and medicine.
Banaś, AM, Bocian-Ostrzycka, KM, Jagusztyn-Krynicka, EK
Critical reviews in microbiology. 2019;(4):433-450
Abstract
The Dsb protein family in prokaryotes catalyzes the generation of disulfide bonds between thiol groups of cysteine residues in nascent proteins, ensuring their proper three-dimensional structure; these bonds are crucial for protein stability and function. The first Dsb protein, Escherichia coli DsbA, was described in 1991. Since then, many details of the bond-formation process have been described through microbiological, biochemical, biophysical and bioinformatics strategies. Research with the model microorganism E. coli and many other bacterial species revealed an enormous diversity of bond-formation mechanisms. Research using Dsb protein engineering has significantly helped to reveal details of the disulfide bond formation. The first part of this review presents the research that led to understanding the mechanism of action of DsbA proteins, which directly transfer their own disulfide into target proteins. The second part concentrates on the mechanism of electron transport through the cell cytoplasmic membrane. Third and lastly, the review discusses the contribution of this research towards new antibacterial agents.
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10.
Wearable and Implantable Soft Bioelectronics Using Two-Dimensional Materials.
Choi, C, Lee, Y, Cho, KW, Koo, JH, Kim, DH
Accounts of chemical research. 2019;(1):73-81
Abstract
Soft bioelectronics intended for application to wearable and implantable biomedical devices have attracted great attention from material scientists, device engineers, and clinicians because of their extremely soft mechanical properties that match with a variety of human organs and tissues, including the brain, heart, skin, eye, muscles, and neurons, as well as their wide diversity in device designs and biomedical functions that can be finely tuned for each specific case of applications. These unique features of the soft bioelectronics have allowed minimal mechanical and biological damage to organs and tissues integrated with bioelectronic devices and reduced side effects including inflammation, skin irritation, and immune responses even after long-term biointegration. These favorable properties for biointegration have enabled long-term monitoring of key biomedical indicators with high signal-to-noise ratio, reliable diagnosis of the patient's health status, and in situ feedback therapy with high treatment efficacy optimized for the requirements of each specific disease model. These advantageous device functions and performances could be maximized by adopting novel high-quality soft nanomaterials, particularly ultrathin two-dimensional (2D) materials, for soft bioelectronics. Two-dimensional materials are emerging material candidates for the channels and electrodes in electronic devices (semiconductors and conductors, respectively). They can also be applied to various biosensors and therapeutic actuators in soft bioelectronics. The ultrathin vertically layered nanostructure, whose layer number can be controlled in the synthesis step, and the horizontally continuous planar molecular structure, which can be found over a large area, have conferred unique mechanical, electrical, and optical properties upon the 2D materials. The atomically thin nanostructure allows mechanical softness and flexibility and high optical transparency of the device, while the large-area continuous thin film structure allows efficient carrier transport within the 2D plane. In addition, the quantum confinement effect in the atomically thin 2D layers introduces interesting optoelectronic properties and superb photodetecting capabilities. When fabricated as soft bioelectronic devices, these interesting and useful material features of the 2D materials enable unconventional device functions in biological and optical sensing, as well as superb performance in electrical and biochemical therapeutic actuations. In this Account, we first summarize the distinctive characteristics of the 2D materials in terms of the mechanical, optical, chemical, electrical, and biomedical aspects and then present application examples of the 2D materials to soft bioelectronic devices based on each aforementioned unique material properties. Among various kinds of 2D materials, we particularly focus on graphene and MoS2. The advantageous material features of graphene and MoS2 include ultrathin thickness, facile functionalization, large surface-to-volume ratio, biocompatibility, superior photoabsorption, and high transparency, which allow the development of high-performance multifunctional soft bioelectronics, such as a wearable glucose patch, a highly sensitive humidity sensor, an ultrathin tactile sensor, a soft neural probe, a soft retinal prosthesis, a smart endoscope, and a cell culture platform. A brief comparison of their characteristics and performances is also provided. Finally, this Account concludes with a future outlook on next-generation soft bioelectronics based on 2D materials.