The Experts below are selected from a list of 705 Experts worldwide ranked by ideXlab platform
Yiming Li - One of the best experts on this subject based on the ideXlab platform.
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Thiol–yne radical reaction mediated site-specific protein labeling via genetic incorporation of an alkynyl-L-lysine analogue
Organic and Biomolecular Chemistry, 2013Co-Authors: Yiming Li, Yitong Li, Yichao HuangAbstract:Three alkyne-containing Pyrrolysine derivatives were synthesized and genetically encoded into proteins by a mutant PylRS–tRNA pair with high efficiencies. With these alkyne handles, site-specific dual labeling of proteins can be achieved via a bioorthogonal thiol–yne ligation reaction.
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thiol yne radical reaction mediated site specific protein labeling via genetic incorporation of an alkynyl l lysine analogue
Organic and Biomolecular Chemistry, 2013Co-Authors: Man Pan, Yitong Li, Yichao Huang, Yiming Li, Qingxiang GuoAbstract:Three alkyne-containing Pyrrolysine derivatives were synthesized and genetically encoded into proteins by a mutant PylRS–tRNA pair with high efficiencies. With these alkyne handles, site-specific dual labeling of proteins can be achieved via a bioorthogonal thiol–yne ligation reaction.
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genetically encoded alkenyl Pyrrolysine analogues for thiol ene reaction mediated site specific protein labeling
Chemical Science, 2012Co-Authors: Maiyun Yang, Xiaoda Song, Yichao Huang, Yiming Li, Peng ChenAbstract:A series of alkene-bearing Pyrrolysine analogues were synthesized and subsequently incorporated into proteins at two sites by a mutant PylRS–tRNA pair with excellent efficiency. This strategy allowed the site-specific labeling of proteins carrying single or double genetically encoded alkene handles via bioorthogonal thiol–ene ligation reactions.
Yichao Huang - One of the best experts on this subject based on the ideXlab platform.
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Thiol–yne radical reaction mediated site-specific protein labeling via genetic incorporation of an alkynyl-L-lysine analogue
Organic and Biomolecular Chemistry, 2013Co-Authors: Yiming Li, Yitong Li, Yichao HuangAbstract:Three alkyne-containing Pyrrolysine derivatives were synthesized and genetically encoded into proteins by a mutant PylRS–tRNA pair with high efficiencies. With these alkyne handles, site-specific dual labeling of proteins can be achieved via a bioorthogonal thiol–yne ligation reaction.
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thiol yne radical reaction mediated site specific protein labeling via genetic incorporation of an alkynyl l lysine analogue
Organic and Biomolecular Chemistry, 2013Co-Authors: Man Pan, Yitong Li, Yichao Huang, Yiming Li, Qingxiang GuoAbstract:Three alkyne-containing Pyrrolysine derivatives were synthesized and genetically encoded into proteins by a mutant PylRS–tRNA pair with high efficiencies. With these alkyne handles, site-specific dual labeling of proteins can be achieved via a bioorthogonal thiol–yne ligation reaction.
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genetically encoded alkenyl Pyrrolysine analogues for thiol ene reaction mediated site specific protein labeling
Chemical Science, 2012Co-Authors: Maiyun Yang, Xiaoda Song, Yichao Huang, Yiming Li, Peng ChenAbstract:A series of alkene-bearing Pyrrolysine analogues were synthesized and subsequently incorporated into proteins at two sites by a mutant PylRS–tRNA pair with excellent efficiency. This strategy allowed the site-specific labeling of proteins carrying single or double genetically encoded alkene handles via bioorthogonal thiol–ene ligation reactions.
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Genetically encoded alkenyl–Pyrrolysine analogues for thiol–ene reaction mediated site-specific protein labeling
Chemical Science, 2012Co-Authors: Maiyun Yang, Xiaoda Song, Yichao Huang, Lei Liu, Peng ChenAbstract:A series of alkene-bearing Pyrrolysine analogues were synthesized and subsequently incorporated into proteins at two sites by a mutant PylRS–tRNA pair with excellent efficiency. This strategy allowed the site-specific labeling of proteins carrying single or double genetically encoded alkene handles via bioorthogonal thiol–ene ligation reactions.
Peng Chen - One of the best experts on this subject based on the ideXlab platform.
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Genetic Code Expansion in Enteric Bacterial Pathogens.
Methods of Molecular Biology, 2018Co-Authors: Huangtao Zheng, Shixian Lin, Peng ChenAbstract:The genetic code expansion strategy has become an elegant method for site-specific incorporation of noncanonical amino acids with diverse functionalities into proteins of interest in bacteria, yeast, mammalian cells, and even animals. This technique allows precise labeling as well as manipulation of a given protein to dissect its physiological and/or pathological roles under living conditions. Here, we demonstrate the extension of a recently emerged Pyrrolysine-based genetic code expansion strategy for encoding noncanonical amino acids into enteric bacterial pathogens.
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Genetically encoded alkenyl–Pyrrolysine analogues for thiol–ene reaction mediated site-specific protein labeling
Chemical Science, 2012Co-Authors: Maiyun Yang, Xiaoda Song, Yichao Huang, Lei Liu, Peng ChenAbstract:A series of alkene-bearing Pyrrolysine analogues were synthesized and subsequently incorporated into proteins at two sites by a mutant PylRS–tRNA pair with excellent efficiency. This strategy allowed the site-specific labeling of proteins carrying single or double genetically encoded alkene handles via bioorthogonal thiol–ene ligation reactions.
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genetically encoded alkenyl Pyrrolysine analogues for thiol ene reaction mediated site specific protein labeling
Chemical Science, 2012Co-Authors: Maiyun Yang, Xiaoda Song, Yichao Huang, Yiming Li, Peng ChenAbstract:A series of alkene-bearing Pyrrolysine analogues were synthesized and subsequently incorporated into proteins at two sites by a mutant PylRS–tRNA pair with excellent efficiency. This strategy allowed the site-specific labeling of proteins carrying single or double genetically encoded alkene handles via bioorthogonal thiol–ene ligation reactions.
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A readily synthesized cyclic Pyrrolysine analogue for site-specific protein “click” labeling
Chemical Communications, 2011Co-Authors: Yanqun Song, Maiyun Yang, Yujie Liang, Jing Wang, Peng ChenAbstract:A concise route was developed for the facile synthesis of a cyclic Pyrrolysine analogue bearing an azide handle. Directed evolution enabled the encoding of this non-natural amino acid in both prokaryotic and eukaryotic cells, which offers a highly efficient approach for the site-specific protein labeling using click chemistry.
Michael K. Chan - One of the best experts on this subject based on the ideXlab platform.
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Pyrrolysine-inspired protein cyclization.
ChemBioChem, 2014Co-Authors: Marianne M. Lee, Liwen Zhang, Tomasz Fekner, Pang-hung Hsu, Bradley S. Heater, Edward J. Behrman, Michael K. ChanAbstract:The Pyrrolysine translational machinery has been extensively explored for the production of recombinant proteins containing a variety of "site-specific" non-canonical amino acids for both in vitro and in vivo biochemical studies. In this study, we report the first use of this technology for the production of branched cyclic proteins with a tadpole-like topology. As a proof of concept, we fused the well-studied RGD peptide to the C terminus of an mCherry reporter protein. Previous studies have shown that cyclization of the RGD peptide enhances its internalization into cells compared to its linear counterpart. The cellular uptake efficiencies of mCherry-cyclo(RGD), mCherry-linear(RGD), and wild-type mCherry determined by flow cytometry follow the trends expected, thereby confirming the feasibility and potential utility of this cyclization approach.
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a click and release Pyrrolysine analogue
ChemBioChem, 2013Co-Authors: Marianne M. Lee, Tomasz Fekner, Lin Wang, Pang-hung Hsu, Tszho Tang, Aurora Hoyin Chan, Michael K. ChanAbstract:What's the catch? A Pyrrolysine analogue bearing a terminal alkyne and an ester functionality can be incorporated into recombinant proteins and render them amenable to capture by the click reaction and subsequent release through ester hydrolysis. The utility of this Pyrrolysine-inspired technology is demonstrated for the identification of SUMOylation sites.
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A Click‐and‐Release Pyrrolysine Analogue
ChemBioChem, 2013Co-Authors: Marianne M. Lee, Tomasz Fekner, Tsz‐ho Tang, Lin Wang, Aurora Ho‐yin Chan, Pang-hung Hsu, Michael K. ChanAbstract:What's the catch? A Pyrrolysine analogue bearing a terminal alkyne and an ester functionality can be incorporated into recombinant proteins and render them amenable to capture by the click reaction and subsequent release through ester hydrolysis. The utility of this Pyrrolysine-inspired technology is demonstrated for the identification of SUMOylation sites.
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The Pyrrolysine translational machinery as a genetic-code expansion tool.
Current Opinion in Chemical Biology, 2011Co-Authors: Tomasz Fekner, Michael K. ChanAbstract:The discovery of Pyrrolysine not only expanded the set of the known proteinogenic amino acids but also revealed unusual features of its encoding mechanism. The engagement of a canonical stop codon and a unique aminoacyl-tRNA synthetase-tRNA pair that can be used to accommodate a broad range of unnatural amino acids while maintaining strict orthogonality in a variety of prokaryotic and eukaryotic expression systems has proven an invaluable combination. Within a few years since its properties were elucidated, the Pyrrolysine translational machinery has become a popular choice for the synthesis of recombinant proteins bearing a wide variety of otherwise hard-to-introduce functional groups. It is also central to the development of new synthetic strategies that rely on stop-codon suppression.
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Pyrrolysine Analogues for Translational Incorporation into Proteins.
ChemInform, 2010Co-Authors: Tomasz Fekner, Michael K. ChanAbstract:Review: design and synthesis of compounds that can act as effective Pyrrolysine mimics, site specifical incorporation into proteins; ca. 60 refs.
Joseph A Krzycki - One of the best experts on this subject based on the ideXlab platform.
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The complete biosynthesis of the genetically encoded amino acid Pyrrolysine from lysine
2016Co-Authors: Marsha A. Gaston, Liwen Zhang, Kari B. Green-church, Joseph A KrzyckiAbstract:Pyrrolysine, the 22nd amino acid to be found in the natural genetic code1–4, is necessary for all known pathways of methane formation from methylamines5,6. The residue is comprised of a methylated pyrroline carboxylate in amide linkage to the ε-amino group of L-lysine2,7,8. The three different methyltransferases that initiate methanogenesis from different methylamines9–11 have genes with an in-frame amber codon12,13 translated as Pyrrolysine2,7,8. E. coli transformed with pylTSBCD from methanogenic Archaea can incorporate endogenously biosynthesized Pyrrolysine into protein14. The decoding of UAG as Pyrrolysine requires pylT1,6 which produces tRNAPyl (also called tRNACUA), and pylS1 encoding a pyrrolysyl-tRNA synthetase4,15,16. The pylBCD genes1 are each required for tRNA-independent Pyrrolysine synthesis14. Pyrrolysine has been the last remaining genetically encoded amino acid with an unknown biosynthetic pathway. Here, we provide genetic and mass spectroscopic evidence for a pylBCD-dependent pathway in which Pyrrolysine arises from two lysines. We show that a new UAG encoded residue, desmethylPyrrolysine, is made from lysine and exogenous D-ornithine in a pylC, then a pylD, dependent process, but is not further converted to Pyrrolysine. These results indicate that th
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A nonPyrrolysine member of the widely distributed trimethylamine methyltransferase family is a glycine betaine methyltransferase.
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Tomislav Ticak, Joseph A Krzycki, Duncan J. Kountz, Kimberly E. Girosky, Donald J. FergusonAbstract:COG5598 comprises a large number of proteins related to MttB, the trimethylamine:corrinoid methyltransferase. MttB has a genetically encoded Pyrrolysine residue proposed essential for catalysis. MttB is the only known trimethylamine methyltransferase, yet the great majority of members of COG5598 lack Pyrrolysine, leaving the activity of these proteins an open question. Here, we describe the function of one of the nonPyrrolysine members of this large protein family. Three nonPyrrolysine MttB homologs are encoded in Desulfitobacterium hafniense, a Gram-positive strict anaerobe present in both the environment and human intestine. D. hafniense was found capable of growth on glycine betaine with electron acceptors such as nitrate or fumarate, producing dimethylglycine and CO2 as products. Examination of the genome revealed genes for tetrahydrofolate-linked oxidation of a methyl group originating from a methylated corrinoid protein, but no obvious means to carry out corrinoid methylation with glycine betaine. DSY3156, encoding one of the nonPyrrolysine MttB homologs, was up-regulated during growth on glycine betaine. The recombinant DSY3156 protein converts glycine betaine and cob(I)alamin to dimethylglycine and methylcobalamin. To our knowledge, DSY3156 is the first glycine betaine:corrinoid methyltransferase described, and a designation of MtgB is proposed. In addition, DSY3157, an adjacently encoded protein, was shown to be a methylcobalamin:tetrahydrofolate methyltransferase and is designated MtgA. Homologs of MtgB are widely distributed, especially in marine bacterioplankton and nitrogen-fixing plant symbionts. They are also found in multiple members of the human microbiome, and may play a beneficial role in trimethylamine homeostasis, which in recent years has been directly tied to human cardiovascular health.
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doi:10.1155/2010/453642 Review Article Selenocysteine, Pyrrolysine, and the Unique Energy
2013Co-Authors: Metabolism Of Methanogenic Archaea, Michael Rother, Joseph A KrzyckiAbstract:License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Methanogenic archaea are a group of strictly anaerobic microorganisms characterized by their strict dependence on the process of methanogenesis for energy conservation. Among the archaea, they are also the only known group synthesizing proteins containing selenocysteine or Pyrrolysine. All but one of the known archaeal Pyrrolysine-containing and all but two of the confirmed archaeal selenocysteine-containing protein are involved in methanogenesis. Synthesis of these proteins proceeds through suppression of translational stop codons but otherwise the two systems are fundamentally different. This paper highlights these differences and summarizes the recent developments in selenocysteine- and Pyrrolysine-related research on archaea and aims to put this knowledge into the context of their unique energy metabolism. 1
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The path of lysine to Pyrrolysine.
Current Opinion in Chemical Biology, 2013Co-Authors: Joseph A KrzyckiAbstract:Pyrrolysine is the 22nd genetically encoded amino acid. For many years, its biosynthesis has been primarily a matter for conjecture. Recently, a pathway for the synthesis of Pyrrolysine from two molecules of lysine was outlined in which a radical SAM enzyme acts as a lysine mutase to generate a methylated ornithine from lysine, which is then ligated to form an amide with the ɛ-amine of a second lysine. Oxidation of the isopeptide gives rise to Pyrrolysine. Mechanisms have been proposed for both the mutase and the ligase, and structures now exist for each, setting the stage for a more detailed understanding of how Pyrrolysine is synthesized and functions in bacteria and archaea.
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PylSn and the Homologous N-terminal Domain of Pyrrolysyl-tRNA Synthetase Bind the tRNA That Is Essential for the Genetic Encoding of Pyrrolysine
Journal of Biological Chemistry, 2012Co-Authors: Ruisheng Jiang, Joseph A KrzyckiAbstract:Pyrrolysine is represented by an amber codon in genes encoding proteins such as the methylamine methyltransferases present in some Archaea and Bacteria. Pyrrolysyl-tRNA synthetase (PylRS) attaches Pyrrolysine to the amber-suppressing tRNA(Pyl). Archaeal PylRS, encoded by pylS, has a catalytic C-terminal domain but an N-terminal region of unknown function and structure. In Bacteria, homologs of the N- and C-terminal regions of archaeal PylRS are respectively encoded by pylSn and pylSc. We show here that wild type PylS from Methanosarcina barkeri and PylSn from Desulfitobacterium hafniense bind tRNA(Pyl) in EMSA with apparent K(d) values of 0.12 and 0.13 μM, respectively. Truncation of the N-terminal region of PylS eliminated detectable tRNA(Pyl) binding as measured by EMSA, but not catalytic activity. A chimeric protein with PylSn fused to the N terminus of truncated PylS regained EMSA-detectable tRNA(Pyl) binding. PylSn did not bind other D. hafniense tRNAs, nor did the competition by the Escherichia coli tRNA pool interfere with tRNA(Pyl) binding. Further indicating the specificity of PylSn interaction with tRNA(Pyl), substitutions of conserved residues in tRNA(Pyl) in the variable loop, D stem, and T stem and loop had significant impact in binding, whereas those having base changes in the acceptor stem or anticodon stem and loop still retained the ability to complex with PylSn. PylSn and the N terminus of PylS comprise the protein superfamily TIGR03129. The members of this family are not similar to any known RNA-binding protein, but our results suggest their common function involves specific binding of tRNA(Pyl).
