Rational Design of Semi‐Synthetic Protein Complexes with the Defined Oligomeric State
31 May 2019-Vol. 4, Iss: 20, pp 6397-6402
About: The article was published on 2019-05-31. It has received 7 citations till now. The article focuses on the topics: Rational design.
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TL;DR: In this article, a new chemical methodology for the rapid synthesis of a diverse set of semi-synthetic protein families, which include protein amphiphiles, facially amphiphilic protein-dendron conjugates, and pH-sensitive protein-Dendron-conjugates, was reported.
Abstract: The design of well-defined monodispersed self-assembling semi-synthetic proteins is emerging as a promising research avenue. These proteins hold great potential to be used as scaffolds for various protein nanotechnology applications. Currently, there are very few chemical methods reported; however, they suffer from elaborate multistep organic synthesis. Herein, we report a new chemical methodology for the rapid synthesis of a diverse set of semi-synthetic protein families, which include protein amphiphiles, facially amphiphilic protein-dendron conjugates, and pH-sensitive protein-dendron conjugates. This chemical method holds great potential to access a wide variety of semi-synthetic proteins in a short time.
5 citations
TL;DR: In this paper, a new chemical methodology for the construction of redox-sensitive protein assemblies using monodisperse facially amphiphilic protein-dendron bioconjugates is presented.
Abstract: The reversible nature of disulfide functionality has been exploited to design intelligent materials such as nanocapsules, micelles, vesicles, inorganic nanoparticles, peptide and nucleic acid nanodevices. Herein, we report a new chemical methodology for the construction redox-sensitive protein assemblies using monodisperse facially amphiphilic protein-dendron bioconjugates. The disulfide functionality is strategically placed between the dendron and protein domains. The custom designed bioconjugates self-assembled into nanoscopic objects of a defined size dictated by the nature of dendron domain. The stimuli-responsive behavior of the protein assemblies is demonstrated using a suitable redox trigger.
2 citations
TL;DR: In this paper, Micelle-assisted protein labeling (MAPLab) technology along with the N-terminal bioconjugation strategy is utilized for the synthesis of a well-defined monodisperse self-assembling semi-synthetic protease.
Abstract: Self-assembly of a monomeric protease to form a multi-subunit protein complex "proteasome" enables targeted protein degradation in living cells. Naturally occurring proteasomes serve as an inspiration and blueprint for the design of artificial protein-based nanoreactors. Here we disclose a general chemical strategy for the design of proteasome-like nanoreactors. Micelle-assisted protein labeling (MAPLab) technology along with the N-terminal bioconjugation strategy is utilized for the synthesis of a well-defined monodisperse self-assembling semi-synthetic protease. The designed protein is programmed to self-assemble into a proteasome-like nanostructure which preserves the functional properties of native protease.
2 citations
TL;DR: In this article, the synthesis of self-assembling artificial proteins (SAPs) using micelle-assisted protein labeling technology (MAPLabTech) has been described by developing a chemical method for site-specific labeling of a surfaceexposed cysteine residue of globular proteins.
Abstract: Self-assembling artificial proteins (SAPs) have gained enormous interest in recent years due to their applications in different fields. Synthesis of well-defined monodisperse SAPs is accomplished predominantly through genetic methods. However, the last decade witnessed the use of few chemical technologies for that purpose. In particular, micelle-assisted protein labeling technology (MAPLabTech) has made huge progress in this area. The first generation MAPLabTech focused on site-specific labeling of the active-site residue of serine proteases to make SAPs. Further, this methodology was exploited for labeling of N-terminal residue of a globular protein to make functional SAPs. In this study, we describe the synthesis of novel SAPs by developing a chemical method for site-specific labeling of a surface-exposed cysteine residue of globular proteins. In addition, we disclose the synthesis of redox- and pH-sensitive SAPs and their systematic self-assembly and dis-assembly studies using complementary biophysical studies. Altogether these studies further expand the scope of MAPLabTech in different fields such as vaccine design, targeted drug delivery, diagnostic imaging, biomaterials, and tissue engineering.
1 citations
TL;DR: The synthesis of novel SAPs are described by developing a chemical method for site‐specific labeling of a surface‐exposed cysteine residue of globular proteins and their systematic self‐assembly and disassembly studies using size‐exclusion chromatography are disclosed.
Abstract: Self‐assembling artificial proteins (SAPs) have gained enormous interest in recent years due to their applications in different fields. Synthesis of well‐defined monodisperse SAPs is accomplished predominantly through genetic methods. However, the last decade has witnessed the use of a few chemical technologies for this purpose. In particular, micelle‐assisted protein labeling technology (MAPLabTech) has made huge progress in this area. The first generation MAPLabTech focused on site‐specific labeling of the active‐site residue of serine proteases to make SAPs. Further, this methodology was exploited for labeling of N‐terminal residue of a globular protein to make functional SAPs. In this study, we describe the synthesis of novel SAPs by developing a chemical method for site‐specific labeling of a surface‐exposed cysteine residue of globular proteins. In addition, we disclose the synthesis of redox‐sensitive SAPs and their systematic self‐assembly and disassembly studies using size‐exclusion chromatography. Altogether these studies further expand the scope of MAPLabTech in different fields such as vaccine design, targeted drug delivery, diagnostic imaging, biomaterials, and tissue engineering.
References
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TL;DR: An understanding of the scope and pattern of the many posttranslational modifications in eukaryotic cells provides insight into the function and dynamics of proteome compositions.
Abstract: The diversity of distinct covalent forms of proteins (the proteome) greatly exceeds the number of proteins predicted by DNA coding capacities owing to directed posttranslational modifications. Enzymes dedicated to such protein modifications include 500 human protein kinases, 150 protein phosphatases, and 500 proteases. The major types of protein covalent modifications, such as phosphorylation, acetylation, glycosylation, methylation, and ubiquitylation, can be classified according to the type of amino acid side chain modified, the category of the modifying enzyme, and the extent of reversibility. Chemical events such as protein splicing, green fluorescent protein maturation, and proteasome autoactivations also represent posttranslational modifications. An understanding of the scope and pattern of the many posttranslational modifications in eukaryotic cells provides insight into the function and dynamics of proteome compositions.
1,214 citations
TL;DR: The chemical synthesis and utility of an active-site directed probe for visualizing dynamics in the expression and function of an entire enzyme family, the serine hydrolases, is described and it is shown that FP-biotin labels these proteins in an activity-dependent manner that can be followed kinetically.
Abstract: With the postgenome era rapidly approaching, new strategies for the functional analysis of proteins are needed. To date, proteomics efforts have primarily been confined to recording variations in protein level rather than activity. The ability to profile classes of proteins on the basis of changes in their activity would greatly accelerate both the assignment of protein function and the identification of potential pharmaceutical targets. Here, we describe the chemical synthesis and utility of an active-site directed probe for visualizing dynamics in the expression and function of an entire enzyme family, the serine hydrolases. By reacting this probe, a biotinylated fluorophosphonate referred to as FP-biotin, with crude tissue extracts, we quickly and with high sensitivity detect numerous serine hydrolases, many of which display tissue-restricted patterns of expression. Additionally, we show that FP-biotin labels these proteins in an activity-dependent manner that can be followed kinetically, offering a powerful means to monitor dynamics simultaneously in both protein function and expression.
987 citations
TL;DR: The accuracy of the method and the number and variety of two-component materials that it makes accessible suggest a route to the construction of functional protein nanomaterials tailored to specific applications.
Abstract: The self-assembly of proteins into highly ordered nanoscale architectures is a hallmark of biological systems. The sophisticated functions of these molecular machines have inspired the development of methods to engineer self-assembling protein nanostructures; however, the design of multi-component protein nanomaterials with high accuracy remains an outstanding challenge. Here we report a computational method for designing protein nanomaterials in which multiple copies of two distinct subunits co-assemble into a specific architecture. We use the method to design five 24-subunit cage-like protein nanomaterials in two distinct symmetric architectures and experimentally demonstrate that their structures are in close agreement with the computational design models. The accuracy of the method and the number and variety of two-component materials that it makes accessible suggest a route to the construction of functional protein nanomaterials tailored to specific applications.
479 citations
TL;DR: A general strategy is described for designing proteins that self assemble into large symmetrical nanomaterials, including molecular cages, filaments, layers, and porous materials, some of which share similar features with natural biological assemblies.
Abstract: A general strategy is described for designing proteins that self assemble into large symmetrical nanomaterials, including molecular cages, filaments, layers, and porous materials. In this strategy, one molecule of protein A, which naturally forms a self-assembling oligomer, An, is fused rigidly to one molecule of protein B, which forms another self-assembling oligomer, Bm. The result is a fusion protein, A-B, which self assembles with other identical copies of itself into a designed nanohedral particle or material, (A-B)p. The strategy is demonstrated through the design, production, and characterization of two fusion proteins: a 49-kDa protein designed to assemble into a cage approximately 15 nm across, and a 44-kDa protein designed to assemble into long filaments approximately 4 nm wide. The strategy opens a way to create a wide variety of potentially useful protein-based materials, some of which share similar features with natural biological assemblies.
415 citations
TL;DR: An analysis of over 5,000 unique atomic structures shows that the quaternary structure of homomers is conserved in over 70% of protein pairs sharing as little as 30% sequence identity, and allows reliable prediction of evolution and assembly of a complex solely from its crystal structure.
Abstract: A homomer is formed by self-interacting copies of a protein unit. This is functionally important, as in allostery, and structurally crucial because mis-assembly of homomers is implicated in disease. Homomers are widespread, with 50-70% of proteins with a known quaternary state assembling into such structures. Despite their prevalence, their role in the evolution of cellular machinery and the potential for their use in the design of new molecular machines, little is known about the mechanisms that drive formation of homomers at the level of evolution and assembly in the cell. Here we present an analysis of over 5,000 unique atomic structures and show that the quaternary structure of homomers is conserved in over 70% of protein pairs sharing as little as 30% sequence identity. Where quaternary structure is not conserved among the members of a protein family, a detailed investigation revealed well-defined evolutionary pathways by which proteins transit between different quaternary structure types. Furthermore, we show by perturbing subunit interfaces within complexes and by mass spectrometry analysis, that the (dis)assembly pathway mimics the evolutionary pathway. These data represent a molecular analogy to Haeckel's evolutionary paradigm of embryonic development, where an intermediate in the assembly of a complex represents a form that appeared in its own evolutionary history. Our model of self-assembly allows reliable prediction of evolution and assembly of a complex solely from its crystal structure.
396 citations