A Genetically Encoded Boronate‐Containing Amino Acid

Brustad, Eric M.; Bushey, Mark L.; Lee, Jae Woo; Groff, Dan; Liu, Wenshe Ray; Schultz, Peter G.

doi:10.1002/anie.200803240

Cited by 158 publications

(175 citation statements)

References 37 publications

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“…The site-specific modification of proteins in this manner avoids the heterogeneous products generated by nonspecific electrophilic reagents and makes possible a form of "protein medicinal chemistry." In addition to these chemistries, borono, iodo, olefinic, and aminophenyl residues have enabled selective modification reactions at the surface of proteins (28).…”

Section: Expanded Genetic Codementioning

confidence: 99%

Beyond the Canonical 20 Amino Acids: Expanding the Genetic Lexicon

Young

Schultz

2010

Journal of Biological Chemistry

309

241

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The ability to genetically encode unnatural amino acids beyond the common 20 has allowed unprecedented control over the chemical structures of recombinantly expressed proteins. Orthogonal aminoacyl-tRNA synthetase/tRNA pairs have been used together with nonsense, rare, or 4-bp codons to incorporate >50 unnatural amino acids into proteins in Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris, and mammalian cell lines. This has allowed the expression of proteins containing amino acids with novel side chains, including fluorophores, post-translational modifications, metal ion chelators, photocaged and photocross-linking moieties, uniquely reactive functional groups, and NMR, IR, and x-ray crystallographic probes. Early MethodologyWith few exceptions, all organisms are restricted to the 20 common amino acid building blocks for the ribosomal biosynthesis of proteins. However, it is clear that proteins require a higher level of chemical complexity for many functions, as evidenced by the frequent use of post-translational modifications and the dependence of many enzymes on cofactors. Thus, the addition of new amino acid building blocks to the genetic code should further expand the range of functions available to proteins and provide powerful new tools for probing protein structure and function both inside and outside of living cells (1). Although recent advances in synthetic and semisynthetic methods have proven very useful for the incorporation of unnatural amino acids into proteins, they are generally limited by low yields and are technically cumbersome in the production of larger proteins. The use of the cellular biosynthetic machinery to introduce novel amino acids abrogates issues relating to scalability and protein size and simplifies the study of modified proteins in living cells.To add a new amino acid to the genetic repertoire, a codon is needed that uniquely specifies that amino acid. The 20 canonical amino acids are encoded by 61 degenerate triplet codons, leaving the remaining three codons (TAG, amber; TAA, ochre; and TGA, opal) to serve as translational stops. Previously, we and others (2) used the redundancy of these "blank" stop codons, together with the ribosomal machinery, to site-specifically incorporate unnatural amino acids into proteins in vitro in response to the amber nonsense codon. This was accomplished by chemically aminoacylating a nonsense suppressor tRNA (tRNA CUA , the amber suppressor tRNA) with the desired unnatural amino acid and adding the aminoacyl-tRNA to a cell-free transcription/translation system along with the gene of interest harboring a TAG mutation at the target site. In the decade that followed, this method was used to site-specifically incorporate a large number of unnatural amino acids with a wide variety of structures into proteins (3, 4). These amino acids were used to probe the roles of specific amino acid side chains and backbone groups in protein folding and stability, catalytic mechanisms, and biomolecular interactions. Later, this approach was extended to th...

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Section: Expanded Genetic Codementioning

confidence: 99%

Beyond the Canonical 20 Amino Acids: Expanding the Genetic Lexicon

Young

Schultz

2010

Journal of Biological Chemistry

309

241

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“…Pioneering work by the group of P. G. Schultz expanded the genetic code via a unique tRNA/aminoacyl-tRNA synthetase pair, resulting in a solid protocol to insert 4-iodophenylalanine [80] and 4-boronophenylalanine [81]. The introduction of 4-iodophenylalanine, for example, opened the gateway for protein modification via Pd-catalyzed reactions [65,82,83].…”

Section: Access To and Derivatization Of (Pseudo)halogenated Aromaticmentioning

confidence: 99%

“…The first example of a Suzuki-Miyaura reaction on protein substrates was reported by Brustad and coworkers via expansion of the genetic code, which allowed selective introduction of a 4-boronophenylalanine residue [83]. Subsequently, the Suzuki-Miyaura reaction induced the coupling of a fluorescent iodoaryl boron-dipyrromethene (bodipy-I) scaffold through use of Pd 2 (dba) 3 complex in an aqueous buffered solution (20 mM EPPS) at 70 • C, though in a moderate conversion (ca.…”

Section: Protein Derivatizationmentioning

confidence: 99%

The Suzuki–Miyaura Cross-Coupling as a Versatile Tool for Peptide Diversification and Cyclization

et al. 2017

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Abstract:The (site-selective) derivatization of amino acids and peptides represents an attractive field with potential applications in the establishment of structure-activity relationships and labeling of bioactive compounds. In this respect, bioorthogonal cross-coupling reactions provide valuable means for ready access to peptide analogues with diversified structure and function. Due to the complex and chiral nature of peptides, mild reaction conditions are preferred; hence, a suitable cross-coupling reaction is required for the chemical modification of these challenging substrates. The Suzuki reaction, involving organoboron species, is appropriate given the stability and environmentally benign nature of these reactants and their amenability to be applied in (partial) aqueous reaction conditions, an expected requirement upon the derivatization of peptides. Concerning the halogenated reaction partner, residues bearing halogen moieties can either be introduced directly as halogenated amino acids during solid-phase peptide synthesis (SPPS) or genetically encoded into larger proteins. A reversed approach building in boron in the peptidic backbone is also possible. Furthermore, based on this complementarity, cyclic peptides can be prepared by halogenation, and borylation of two amino acid side chains present within the same peptidic substrate. Here, the Suzuki-Miyaura reaction is a tool to induce the desired cyclization. In this review, we discuss diverse amino acid and peptide-based applications explored by means of this extremely versatile cross-coupling reaction. With the advent of peptide-based drugs, versatile bioorthogonal conversions on these substrates have become highly valuable.

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“…14 UAA*s ( Fig. 2) that are successfully applied to the biosensor design include 3,4-dihydroxy-L-phenylalanine (L-DOPA; metal ion sensor), 10,45 2-amino-3-(8-hydroxyquinolin-5-yl)propanoic acid (HqAla; metal ion sensor), 13 para-borono-L-phenylalanine (pBoF; H 2 O 2 sensor), 11,44 para-azido-L-phenylalanine (pAzF; H 2 S sensor), 12,45 and o-nitrobenzyl-O-tyrosine (ONBY, light-responsive FP). 14,46 49 HqAla, 13 and bipyridyl-L-alanine (BpA), 50 into fluorescent proteins.…”

Section: Review Molecular Biosystemsmentioning

confidence: 99%