2.2 CuAAC in Peptidomimetics and Protein Mimics

The tremendous recent developments in click chemistry, including the impressive developments of strain-promoted cycloaddition reagents, all started with the copper-catalyzed azide–alkyne cycloaddition (CuAAC) reaction conceived by Meldal et al. and Sharpless et al. This led to a revolution of extremely important applications in the chemical, biological, medical, and materials sciences. It is fair to state that, especially in the synthesis of multifunctional and complex small-to-large biomolecular constructs, CuAAC has been indispensable. This has been particularly evident in the area of peptides, peptidomimetics, and protein mimics. These biomolecules play key roles in the various peptide–peptide, peptide–protein, and protein–protein interactions that are involved in many diseases and disorders, and peptide-based therapeutics can be important in this context. However, it is often important to improve the bioactivity and overall stability, and modulate the spatial structure, of peptide-based therapeutics. The incorporation of the 1,4-disubstituted 1,2,3-triazole moiety as a non-native structural element using CuAAC is explored in this chapter. The resulting incorporated triazole moiety can lead to structural surrogates of the amide bond and disulfide bond. As a consequence, CuAAC can be utilized toward introducing conformational constraints and stabilizing secondary structures of α-helices, β-sheets/turns, or loop-like structures. In addition, CuAAC can be used to combine various peptide sequences with molecular scaffolds to develop protein mimics that can find applications as synthetic vaccines and antibodies.

Identification of a Mimicry of the Protein that Potentially Isolates the Mutated p53 Material and Prevents Further Protein Accumulation

The team first screened a set of protein mimics originally designed to target Alzheimer's disease and type 2 diabetes. The results identify a mimicry of the protein that potentially isolates the mutated p53 material and prevents further protein accumulation. The researchers then showed that segregation of mutated p53 grains by protein mimicking restored the suppressive function of the p53 tumor, leading to the death of a wide range of cancer cells. Importantly, protein mimicry therapy effectively reduces tumors that contain mutated p53 while showing no significant toxins for healthy tissue, resulting in significantly longer survival. "As the prevalence of cancer increases worldwide, there is an urgent need for new cancer therapies to complement or replace existing therapies," said the study's lead author. Here we show the first successful use of a small molecule amyloid inhibitor as an anticancer agent. We believe that this will have a far-reaching impact, as it effectively bridges the gap between amyloid disease and cancer and is the basis for passing on information approaches in the design of new and robust cancer mutation therapies for the p53 mutation. Keywords: Cancer; Cells; Tissues; Tumors; Prevention; Prognosis; Diagnosis; Imaging; Screening, Treatment; Management

Chaperones drive in vitro evolution of uracil glycosylase towards misfolded states

Evolution is driven by random mutations, whose fitness outcome is tested over time. In vitro evolution of a library of a randomly mutated protein mimics this process, however, on a short time scale, driven by a specific outcome (such as binding to a bait). Here, we used directed in vitro evolution to investigate the role of molecular chaperones in curbing promiscuity in favor of specificity of protein-protein interactions. Using yeast surface display, we generated a random library of the E. coli protein Uracil glycosylase (UNG), and selected it against various baits. Those included the purified chaperones GroEL, DnaK+DnaJ+ATP, or total protein extracts from WT or delta DnaK+DnaJ cells. We show that in-vitro evolution differs from natural evolution in cells, both physically and thermodynamically. We found that chaperones, whether purified or as part of the protein extract, select for and thus enrich uracil glycosylase (UNG) misfolded species during this in vitro evolution process. In a more general context, our results show that chaperones purge promiscuous misfolded clones from the system, and thereby avoiding their detrimental effects, such as forming wrong interactions with other macromolecules, including proteins, which can harm proteostasis.

Targeting the Prion-Like Aggregation of Mutant p53 to Combat Cancer

The team first screened a set of protein mimics originally designed to target Alzheimer's disease and type 2 diabetes. The results identify a mimicry of the protein that potentially isolates the mutated p53 material and prevents further protein accumulation. The researchers then showed that segregation of mutated p53 grains by protein mimicking restored the suppressive function of the p53 tumor, leading to the death of a wide range of cancer cells. Importantly, protein mimicry therapy effectively reduces tumors that contain mutated p53 while showing no significant toxins for healthy tissue, resulting in significantly longer survival. "As the prevalence of cancer increases worldwide, there is an urgent need for new cancer therapies to complement or replace existing therapies," said the study's lead author. Here we show the first successful use of a small molecule amyloid inhibitor as an anticancer agent. We believe that this will have a far-reaching impact, as it effectively bridges the gap between amyloid disease and cancer and is the basis for passing on information approaches in the design of new and robust cancer mutation therapies for the p53 mutation. Keywords: Cancer; Cells; Tissues, Tumors; Prevention, Prognosis; Diagnosis; Imaging; Screening; Treatment; Management

Protein-Based Systems for Topical Antibacterial Therapy

Recently, proteins are gaining attention as potential materials for antibacterial therapy. Proteins possess beneficial properties such as biocompatibility, biodegradability, low immunogenic response, ability to control drug release, and can act as protein-mimics in wound healing. Different plant- and animal-derived proteins can be developed into formulations (films, hydrogels, scaffolds, mats) for topical antibacterial therapy. The application areas for topical antibacterial therapy can be wide including bacterial infections in the skin (e.g., acne, wounds), eyelids, mouth, lips, etc. One of the major challenges of the healthcare system is chronic wound infections. Conventional treatment strategies for topical antibacterial therapy of infected wounds are inadequate, and the development of newer and optimized formulations is warranted. Therefore, this review focuses on recent advances in protein-based systems for topical antibacterial therapy in infected wounds. The opportunities and challenges of such protein-based systems along with their future prospects are discussed.

A robust polyfunctional Pd(II)-based magnetic amphiphilic nanocatalyst for the Suzuki–Miyaura coupling reaction

Herein, a robust Pd(II)-based polyfunctional magnetic amphiphilic artificial metalloenzyme was prepared by anchoring a Pd(2,2′-dipyridylamine)Cl2 bearing hydrophilic monomethyl ether poly(ethylene glycol) (mPEG) chains on the surface of amino-functionalized silica-coated magnetic nanoparticles. The 2,2′-dipyridylamine (dpa) has shown excellent complexation properties for Pd(II) and it could be easily anchored onto functionalized magnetic support by the bridging nitrogen atom. Moreover, the bridging nitrogen atom at the proximity of Pd(II) catalytic center could play an important role in dynamic suppramolecular interactions with substrates. The leaching, air and moisture resistant [Pd(dpa)Cl2] complex endow the dynamic and robust structure to the designed artificial enzyme. Moreover, the water dispersibility of designed artificial metalloenzyme raised from mPEG chains and the magnetic nanoparticles core which could function as protein mimics endow it other necessary characters of artificial enzymes. The prepared artificial metalloenzyme displayed remarkable activity in Suzuki–Miyaura cross-coupling reaction employing low-palladium loading under mild conditions, with the exceptionally high turnover frequency, clean reaction profile, easy work-up procedure, good to excellent products yields and short reaction times. The designed air- and moisture-stable artificial metalloenzyme could recycle more than fifteen times with easy separation procedure in aqueous solution under aerobic conditions without any noticeable loss in activity.

Viral mimicry of p65/RelA transactivation domain to inhibit NF-κB activation

ABSTRACTThe evolutionary arms race between hosts and their viruses drove the evolution of complex immune systems in mammals and sophisticated immune evasion mechanisms by viruses. Mammalian antiviral defences require sensing of virus infection and stimulation of the expression of interferons and cytokines via the activation of NF-κB and other immune signalling pathways. Viruses antagonise these host antiviral defences by interfering with immune sensing and signal transduction and blocking the actions of interferons and cytokines. Here we show that a viral protein mimics the transcription activation domain of p65, the transcriptionally active subunit of NF-κB. The C terminus of vaccinia virus (VACV) protein F14 (residues 51-73) activates transcription when fused to a DNA-binding domain-containing protein and associates with NF-κB co-activator CBP, disrupting its interaction with p65. Consequently, F14 diminishes CBP-mediated acetylation of p65 and the downstream recruitment of RNA polymerase II processivity factor BRD4 to the promoter of NF-κB-responsive gene CXCL10, thereby inhibiting the expression and secretion of CXCL10 upon stimulation with TNF-α. A VACV strain engineered to lack F14 caused reduced lesions in an intradermal model of infection, showing that F14 contributes to virulence. Our results uncover a mechanism by which viruses disarm the antiviral defences through molecular mimicry of a conserved protein of the host’s immune system.

A Cysteine Selenosulfide Redox Switch for Protein Chemical Synthesis

<div>We describe a cyclic selenosulfide derivative of cysteine called SetCys that enables to perform protein chemical synthesis under redox-control.</div><div>Native chemical ligation or SEA-mediated ligation with SetCys peptide segments proceeds in a traceless manner and involves the cleavage of a carbon-nitrogen bond in situ.</div><div>The manuscript describes detailed mechanistic investigations of SetCys redox-switch and its application to the production of biologically active cyclic protein mimics of hepatocyte growth factor, the high-affinity ligand of MET tyrosine kinase receptor.<br></div><div><br></div>