Abstract:Native chemical ligation has enabled the chemical synthesis of proteins for a wide variety of applications (e.g., mirror-image proteins). However, inefficiencies of this chemoselective ligation in the context of large or otherwise challenging protein targets can limit the practical scope of chemical protein synthesis. In this review, we focus on recent developments aimed at enhancing and expanding native chemical ligation for challenging protein syntheses. Chemical auxiliaries, use of selenium chemistry, and t… Show more
“…NCL is classically run at mM peptide concentrations and typically displays second-order rate constants of 0.3–4 M −1 s −1 32 , 33 . This reaction does not proceed at µM peptide concentrations unless it is assisted by a biomolecular template or other means 34 , 35 . Fig.…”
The modification of protein electrostatics by phosphorylation is a mechanism used by cells to promote the association of proteins with other biomolecules. In this work, we show that introducing negatively charged phosphoserines in a reactant is a powerful means for directing and accelerating the chemical modification of proteins equipped with oppositely charged arginines. While the extra charged amino acid residues induce no detectable affinity between the reactants, they bring site-selectivity to a reaction that is otherwise devoid of such a property. They also enable rate accelerations of four orders of magnitude in some cases, thereby permitting chemical processes to proceed at the protein level in the low micromolar range, using reactions that are normally too slow to be useful in such dilute conditions.
“…NCL is classically run at mM peptide concentrations and typically displays second-order rate constants of 0.3–4 M −1 s −1 32 , 33 . This reaction does not proceed at µM peptide concentrations unless it is assisted by a biomolecular template or other means 34 , 35 . Fig.…”
The modification of protein electrostatics by phosphorylation is a mechanism used by cells to promote the association of proteins with other biomolecules. In this work, we show that introducing negatively charged phosphoserines in a reactant is a powerful means for directing and accelerating the chemical modification of proteins equipped with oppositely charged arginines. While the extra charged amino acid residues induce no detectable affinity between the reactants, they bring site-selectivity to a reaction that is otherwise devoid of such a property. They also enable rate accelerations of four orders of magnitude in some cases, thereby permitting chemical processes to proceed at the protein level in the low micromolar range, using reactions that are normally too slow to be useful in such dilute conditions.
“…Another more original linkage method based on the native chemical ligation (NCL) [48] was applied to AAZTA derivatives by Hawala et al [49]. This technique allowed the formation of an amide bond between a N-terminal cysteinyl peptide and a pentanoic acid-AAZTA derivative, preactivated as a sodium 2-mercaptoethanesulfonate thioester (Scheme 7).…”
Section: Aazta Modulations To Obtain Bifunctional Chelating Agents (Bca)mentioning
With the development of 68Ga and 177Lu radiochemistry, theranostic approaches in modern nuclear medicine enabling patient-centered personalized medicine applications have been growing in the last decade. In conjunction with the search for new relevant molecular targets, the design of innovative chelating agents to easily form stable complexes with various radiometals for theranostic applications has gained evident momentum. Initially conceived for magnetic resonance imaging applications, the chelating agent AAZTA features a mesocyclic seven-membered diazepane ring, conferring some of the properties of both acyclic and macrocyclic chelating agents. Described in the early 2000s, AAZTA and its derivatives exhibited interesting properties once complexed with metals and radiometals, combining a fast kinetic of formation with a slow kinetic of dissociation. Importantly, the extremely short coordination reaction times allowed by AAZTA derivatives were particularly suitable for short half-life radioelements (i.e., 68Ga). In view of these particular characteristics, the scope of this review is to provide a survey on the design, synthesis, and applications in the nuclear medicine/radiopharmacy field of AAZTA-derived chelators.
“…23 Since most of the immune coreceptors contain a nonstructural cytoplasmic domain with a relatively short AA sequence, this protein fragment-based AP-MS analysis is expected to be broadly applied for profiling signaling complexes in immunological systems. In addition, as full length protein synthesis can be achieved by native chemical ligation (NCL), 50 expressed protein ligation (EPL), 51 and the latest automated flow chemistry, 52 we would expect their growing application for profiling PTM-dependent protein complexes. Integrated Sample Preparation for AP-MS.…”
“…In the aforementioned trifunctional probe-based methods, ligand proteins are typically prepared through recombinant expression or purification, making it hard to site-specifically introduce the PTMs of interest into the protein ligand. As mentioned above, chemical protein synthesis offers site-specific installation of desired PTMs and unnatural amino acids (UAAs) which carry diazirine and other functional groups. , The combination with NCL and EPL technologies further empowers its application to a full-length protein rather than the PTM-containing fragments. For example, the diubiquitins-based photoaffinity probes linked via Lys48 or Lys63 have been chemically synthesized for photo-cross-linking and AP-MS analysis of linkage-specific ubiquitin-binding proteins .…”
Section: Ligand-directed Cross-linking
For Protein Complex Profilingmentioning
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