Convergent diversity-oriented side-chain macrocyclization scan for unprotected polypeptides

Here we describe a general synthetic platform for side-chain macrocyclization of an unprotected peptide library based on the S[subscript N]Ar reaction between cysteine thiolates and a new generation of highly reactive perfluoroaromatic small molecule linkers. This strategy enabled us to simultaneously “scan” two cysteine residues positioned from i, i + 1 to i, i + 14 sites in a polypeptide, producing 98 macrocyclic products from reactions of 14 peptides with 7 linkers. A complementary reverse strategy was developed; cysteine residues within the polypeptide were first modified with non-bridging perfluoroaryl moieties and then commercially available dithiol linkers were used for macrocyclization. The highly convergent, site-independent, and modular nature of these two strategies coupled with the unique chemoselectivity of a S[subscript N]Ar transformation allows for the rapid diversity-oriented synthesis of hybrid macrocyclic peptide libraries with varied chemical and structural complexities.National Institutes of Health (U.S.) (GM101762)National Institutes of Health (U.S.) (GM046059)MIT Faculty Start-up FundSontag Foundation (Distinguished Scientist Award)Deshpande Center for Technological InnovationMassachusetts Institute of Technology (Charles E. Reed Faculty Initiative Fund)Damon Runyon Cancer Research Foundatio

Sterically Unprotected Nucleophilic Boron Cluster Reagents

A cornerstone of modern synthetic chemistry rests on the ability to manipulate the reactivity of a carbon center by rendering it either electrophilic or nucleophilic. However, accessing a similar reactivity spectrum with boron-based reagents has been significantly more challenging. While classical nucleophilic carbon-based reagents normally do not require steric protection, readily accessible, unprotected boron-based nucleophiles have not yet been realized. Herein, we demonstrate that the bench stable closo-hexaborate cluster anion can engage in a nucleophilic substitution reaction with a wide array of organic and main group electrophiles. The resulting molecules containing B‒C bonds can be further converted to tricoordinate boron species widely used in organic synthesis

Rapid Flow-Based Peptide Synthesis

A flow-based solid-phase peptide synthesis methodology that enables the incorporation of an amino acid residue every 1.8 min under automatic control or every 3 min under manual control is described. This is accomplished by passing a stream of reagent through a heat exchanger into a low volume, low backpressure reaction vessel, and through a UV detector. These features enable continuous delivery of heated solvents and reagents to the solid support at high flow rate, thereby maintaining maximal concentration of reagents in the reaction vessel, quickly exchanging reagents, and eliminating the need to rapidly heat reagents after they have been added to the vessel. The UV detector enables continuous monitoring of the process. To demonstrate the broad applicability and reliability of this method, it was employed in the total synthesis of a small protein, as well as dozens of peptides. The quality of the material obtained with this method is comparable to that for traditional batch methods, and, in all cases, the desired material was readily purifiable by RP-HPLC. The application of this method to the synthesis of the 113-residue Bacillus amyloliquefaciens RNase and the 130-residue DARPin pE59 is described in the accompanying manuscript.MIT Faculty Start-up FundMassachusetts Institute of Technology (Charles E. Reed Faculty Initiative Fund)Deshpande Center for Technological InnovationDamon Runyon-Rachleff (Innovation Award)Sontag Foundation (Distinguished Scientist Award)C. P. Chu and Y. Lai FellowshipDaniel S. Kemp Summer FellowshipNational Institute of General Medical Sciences (U.S.). Biotechnology Training Program (Grant 5T32GM008334-25)National Institutes of Health (U.S.) (Fellowship F32GM101762

Ultra-fast Au(III)-mediated Arylation of Cysteine

Through mechanistic work and rational design, we have developed the fastest organometallic abiotic Cys bioconjugation. As a result, the developed organometallic Au(III) bioconjugation reagents enable selective labeling of Cys moieties down to pM concentrations and allow for the rapid construction of complex heterostructures from peptides, proteins, and oligonucleo-tides. This work showcases how organometallic chemistry can be interfaced with biomolecules and lead to the range of reac-tivities that are largely unmatched by classical organic chemistry tools

A Super-Oxidized Radical Cationic Icosahedral Boron Cluster

While the icosahedral closo-[B₁₂H₁₂]²⁻ cluster does not display reversible electrochemical behavior, perfunctionalization of this species via substitution of all 12 B–H vertices with alkoxy or benzyloxy (OR) substituents engenders reversible redox chemistry, providing access to clusters in the dianionic, monoanionic, and neutral forms. Here, we evaluated the electrochemical behavior of the electron-rich B₁₂(O-3-methylbutyl)₁₂ (1) cluster and discovered that a new reversible redox event that gives rise to a fourth electronic state is accessible through one-electron oxidation of the neutral species. Chemical oxidation of 1 with [N(2,4-Br₂C₆H₃)₃]·⁺ afforded the isolable [1]·⁺ cluster, which is the first example of an open-shell cationic B₁₂ cluster in which the unpaired electron is proposed to be delocalized throughout the boron cluster core. The oxidation of 1 is also chemically reversible, where treatment of [1]·⁺ with ferrocene resulted in its reduction back to 1. The identity of [1]·⁺ is supported by EPR, UV–vis, multinuclear NMR (¹H, ¹¹B), and X-ray photoelectron spectroscopic characterization

A Super-Oxidized Radical Cationic Icosahedral Boron Cluster

While the icosahedral closo-[B₁₂H₁₂]²⁻ cluster does not display reversible electrochemical behavior, perfunctionalization of this species via substitution of all 12 B–H vertices with alkoxy or benzyloxy (OR) substituents engenders reversible redox chemistry, providing access to clusters in the dianionic, monoanionic, and neutral forms. Here, we evaluated the electrochemical behavior of the electron-rich B₁₂(O-3-methylbutyl)₁₂ (1) cluster and discovered that a new reversible redox event that gives rise to a fourth electronic state is accessible through one-electron oxidation of the neutral species. Chemical oxidation of 1 with [N(2,4-Br₂C₆H₃)₃]·⁺ afforded the isolable [1]·⁺ cluster, which is the first example of an open-shell cationic B₁₂ cluster in which the unpaired electron is proposed to be delocalized throughout the boron cluster core. The oxidation of 1 is also chemically reversible, where treatment of [1]·⁺ with ferrocene resulted in its reduction back to 1. The identity of [1]·⁺ is supported by EPR, UV–vis, multinuclear NMR (¹H, ¹¹B), and X-ray photoelectron spectroscopic characterization

Organometallic palladium reagents for cysteine bioconjugation

Reactions based on transition metals have found wide use in organic synthesis, in particular for the functionalization of small molecules. However, there are very few reports of using transition-metal-based reactions to modify complex biomolecules, which is due to the need for stringent reaction conditions (for example, aqueous media, low temperature and mild pH) and the existence of multiple reactive functional groups found in biomolecules. Here we report that palladium(II) complexes can be used for efficient and highly selective cysteine conjugation (bioconjugation) reactions that are rapid and robust under a range of bio-compatible reaction conditions. The straightforward synthesis of the palladium reagents from diverse and easily accessible aryl halide and trifluoromethanesulfonate precursors makes the method highly practical, providing access to a large structural space for protein modification. The resulting aryl bioconjugates are stable towards acids, bases, oxidants and external thiol nucleophiles. The broad utility of the bioconjugation platform was further corroborated by the synthesis of new classes of stapled peptides and antibody–drug conjugates. These palladium complexes show potential as benchtop reagents for diverse bioconjugation applications.National Institutes of Health (U.S.) (GM-58160)National Institutes of Health (U.S.) (GM-101762)MIT Faculty Start-up FundDamon Runyon Cancer Research FoundationSontag Foundation (Distinguished Scientist Award)Massachusetts Institute of Technology. Dept. of Chemistry (George Buchi Research Fellowship)David H. Koch Institute for Integrative Cancer Research at MIT (Graduate Fellowship in Cancer Research

Enzyme-Catalyzed Macrocyclization of Long Unprotected Peptides

A glutathione S-transferase (GST) catalyzed macrocyclization reaction for peptides up to 40 amino acids in length is reported. GST catalyzes the selective SNAr reaction between an N-terminal glutathione (GSH, γ-Glu-Cys-Gly) tag and a C-terminal perfluoroaryl-modified cysteine on the same polypeptide chain. Cyclic peptides ranging from 9 to 24 residues were quantitatively produced within 2 h in aqueous pH = 8 buffer at room temperature. The reaction was highly selective for cyclization at the GSH tag, enabling the combination of GST-catalyzed ligation with native chemical ligation to generate a large 40-residue peptide macrocycle.Massachusetts Institute of Technology (MIT startup funds)National Institutes of Health (U.S.) (grant GM101762)Damon Runyon Cancer Research Foundation (Award)Sontag Foundation (Distinguished Scientist Award)Amgen Inc. (Summer Graduate Research Fellowship

Visible-Light-Induced Olefin Activation Using 3D Aromatic Boron-Rich Cluster Photooxidants

We report a discovery that perfunctionalized icosahedral dodecaborate clusters of the type B_(12)(OCH_2Ar)_(12) (Ar = Ph or C_6F_5) can undergo photo-excitation with visible light, leading to a new class of metal-free photooxidants. Excitation in these species occurs as a result of the charge transfer between low-lying orbitals located on the benzyl substituents and an unoccupied orbital delocalized throughout the boron cluster core. Here we show how these species, photo-excited with a benchtop blue LED source, can exhibit excited-state reduction potentials as high as 3 V and can participate in electron-transfer processes with a broad range of styrene monomers, initiating their polymerization. Initiation is observed in cases of both electron-rich and electron-deficient styrene monomers at cluster loadings as low as 0.005 mol%. Furthermore, photo-excitation of B_(12)(OCH_2C_6F_5)_(12) in the presence of a less activated olefin such as isobutylene results in the production of highly branched poly(isobutylene). This work introduces a new class of air-stable, metal-free photo-redox reagents capable of mediating chemical transformations