Modern Extensions of Native Chemical Ligation for Chemical Protein Synthesis

  • Lara R. MalinsEmail author
  • Richard J. PayneEmail author
Part of the Topics in Current Chemistry book series (TOPCURRCHEM, volume 362)


Over the past 20 years, native chemical ligation has facilitated the synthesis of numerous complex peptide and protein targets, with and without post-translational modifications, as well as the design and construction of a variety of engineered protein variants. This powerful methodology has also served as a platform for the development of related chemoselective ligation technologies which have greatly expanded the scope and flexibility of ligation chemistry. This chapter details a number of important extensions of the original native chemical ligation manifold, with particular focus on the application of new methods in the total chemical synthesis of proteins. Topics covered include the development of auxiliary-based ligation methods, the post-ligation manipulation of Cys residues, and the synthesis and utility of unnatural amino acid building blocks (bearing reactive thiol or selenol functionalities) in chemoselective ligation chemistry. Contemporary applications of these techniques to the total chemical synthesis of peptides and proteins are described.


Deselenization Desulfurization Native chemical ligation Post-translational modifications Protein synthesis 


  1. 1.
    Walsh CT, Garneau-Tsodikova S, Gatto GJ Jr (2005) Protein posttranslational modifications: the chemistry of proteome diversifications. Angew Chem Int Ed 44:7342–7372Google Scholar
  2. 2.
    Walsh C (2006) Posttranslational modification of proteins: expanding nature’s inventory. Roberts and Co Publishers, Englewood, ColoGoogle Scholar
  3. 3.
    Dawson PE, Muir TW, Clark-Lewis I, Kent SBH (1994) Synthesis of proteins by native chemical ligation. Science 266:776–779Google Scholar
  4. 4.
    Wieland T, Bokelmann E, Bauer L, Lang HU, Lau H (1953) Über peptidsynthesen. 8. Mitteilung bildung von S-haltigen peptiden durch intramolekulare wanderung von aminoacylresten. Justus Liebigs Ann Chem 583:129–149Google Scholar
  5. 5.
    Merrifield RB (1963) Solid phase peptide synthesis. I. The synthesis of a tetrapeptide. J Am Chem Soc 85:2149–2154Google Scholar
  6. 6.
    Kemp DS, Leung S-L, Kerkman DJ (1981) Models that demonstrate peptide bond formation by prior thiol capture. I. Capture by disulfide formation. Tetrahedron Lett 22:181–184Google Scholar
  7. 7.
    Kemp DS, Kerkman DJ (1981) Models that demonstrate peptide bond formation by prior thiol capture. II. Capture by organomercury derivatives. Tetrahedron Lett 22:185–186Google Scholar
  8. 8.
    Fotouhi N, Galakatos NG, Kemp DS (1989) Peptide synthesis by prior thiol capture. 6. Rates of the disulfide-bond-forming capture reaction and demonstration of the overall strategy by synthesis of the C-terminal 29-peptide sequence of BPTI. J Org Chem 54:2803–2817Google Scholar
  9. 9.
    Liu C-F, Tam JP (1994) Chemical ligation approach to form a peptide bond between unprotected peptide segments. Concept and model study. J Am Chem Soc 116:4149–4153Google Scholar
  10. 10.
    Liu C-F, Tam JP (1994) Peptide segment ligation strategy without use of protecting groups. Proc Natl Acad Sci U S A 91:6584–6588Google Scholar
  11. 11.
    Schnoelzer M, Kent SBH (1992) Constructing proteins by dovetailing unprotected synthetic peptides: backbone-engineered HIV protease. Science 256:221–225Google Scholar
  12. 12.
    Schnoelzer M, Alewood P, Jones A, Alewood D, Kent SBH (1992) In situ neutralization in Boc-chemistry solid phase peptide synthesis. Rapid, high yield assembly of difficult sequences. Int J Pept Protein Res 40:180–193Google Scholar
  13. 13.
    Hackeng TM, Griffin JH, Dawson PE (1999) Protein synthesis by native chemical ligation: expanded scope by using straightforward methodology. Proc Natl Acad Sci U S A 96:10068–10073Google Scholar
  14. 14.
    Camarero J, Adeva A, Muir T (2000) 3-Thiopropionic acid as a highly versatile multidetachable thioester resin linker. Lett Pept Sci 7:17–21Google Scholar
  15. 15.
    Dawson PE, Churchill MJ, Ghadiri MR, Kent SBH (1997) Modulation of reactivity in native chemical ligation through the use of thiol additives. J Am Chem Soc 119:4325–4329Google Scholar
  16. 16.
    Johnson ECB, Kent SBH (2006) Insights into the mechanism and catalysis of the native chemical ligation reaction. J Am Chem Soc 128:6640–6646Google Scholar
  17. 17.
    Evans TC Jr, Benner J, Xu MQ (1998) Semisynthesis of cytotoxic proteins using a modified protein splicing element. Protein Sci 7:2256–2264Google Scholar
  18. 18.
    Muir TW (2003) Semisynthesis of proteins by expressed protein ligation. Annu Rev Biochem 72:249–289Google Scholar
  19. 19.
    Pollock SB, Kent SB (2011) An investigation into the origin of the dramatically reduced reactivity of peptide-prolyl-thioesters in native chemical ligation. Chem Commun 47:2342–2344Google Scholar
  20. 20.
    Kent SB (2009) Total chemical synthesis of proteins. Chem Soc Rev 38:338–351Google Scholar
  21. 21.
    Payne RJ, Wong CH (2010) Advances in chemical ligation strategies for the synthesis of glycopeptides and glycoproteins. Chem Commun 46:21–43Google Scholar
  22. 22.
    Raibaut L, Ollivier N, Melnyk O (2012) Sequential native peptide ligation strategies for total chemical protein synthesis. Chem Soc Rev 41:7001–7015Google Scholar
  23. 23.
    Macmillan D (2006) Evolving strategies for protein synthesis converge on native chemical ligation. Angew Chem Int Ed 45:7668–7672Google Scholar
  24. 24.
    Hackenberger CPR, Schwarzer D (2008) Chemoselective ligation and modification strategies for peptides and proteins. Angew Chem Int Ed 47:10030–10074Google Scholar
  25. 25.
    Unverzagt C, Kajihara Y (2013) Chemical assembly of N-glycoproteins: a refined toolbox to address a ubiquitous posttranslational modification. Chem Soc Rev 42:4408–4420Google Scholar
  26. 26.
    Haase C, Seitz O (2008) Extending the scope of native chemical peptide coupling. Angew Chem Int Ed 47:1553–1556Google Scholar
  27. 27.
    Dirksen A, Dawson PE (2008) Expanding the scope of chemoselective peptide ligations in chemical biology. Curr Opin Chem Biol 12:760–766Google Scholar
  28. 28.
    Gamblin DP, Scanlan EM, Davis BG (2009) Glycoprotein synthesis: an update. Chem Rev 109:131–163Google Scholar
  29. 29.
    Okamoto R, Mandal K, Ling M, Luster AD, Kajihara Y, Kent SBH (2014) Total chemical synthesis and biological activities of glycosylated and non-glycosylated forms of the chemokines CCL1 and Ser-CCL1. Angew Chem Int Ed 53:5188–5193Google Scholar
  30. 30.
    Bang D, Kent SB (2004) A one-pot total synthesis of crambin. Angew Chem Int Ed 43:2534–2538Google Scholar
  31. 31.
    Blanco-Canosa JB, Dawson PE (2008) An efficient Fmoc-SPPS approach for the generation of thioester peptide precursors for use in native chemical ligation. Angew Chem Int Ed 47:6851–6855Google Scholar
  32. 32.
    Okamoto R, Mandal K, Sawaya MR, Kajihara Y, Yeates TO, Kent SB (2014) (Quasi-)racemic X-ray structures of glycosylated and non-glycosylated forms of the chemokine Ser-CCL1 prepared by total chemical synthesis. Angew Chem Int Ed 53:5194–5198Google Scholar
  33. 33.
    UniprotKB/TrEMBL Protein Database Release 2014_07 Statistics (2014) Accessed 1 Sept 2014
  34. 34.
    Canne LE, Bark SJ, Kent SBH (1996) Extending the applicability of native chemical ligation. J Am Chem Soc 118:5891–5896Google Scholar
  35. 35.
    Marinzi C, Bark SJ, Offer J, Dawson PE (2001) A new scaffold for amide ligation. Bioorg Med Chem 9:2323–2328Google Scholar
  36. 36.
    Botti P, Carrasco MR, Kent SBH (2001) Native chemical ligation using removable Nα-(1-phenyl-2-mercaptoethyl) auxiliaries. Tetrahedron Lett 42:1831–1833Google Scholar
  37. 37.
    Low DW, Hill MG, Carrasco MR, Kent SB, Botti P (2001) Total synthesis of cytochrome b562 by native chemical ligation using a removable auxiliary. Proc Natl Acad Sci U S A 98:6554–6559Google Scholar
  38. 38.
    Kawakami T, Aimoto S (2003) A photoremovable ligation auxiliary for use in polypeptide synthesis. Tetrahedron Lett 44:6059–6061Google Scholar
  39. 39.
    Marinzi C, Offer J, Longhi R, Dawson PE (2004) An o-nitrobenzyl scaffold for peptide ligation: synthesis and applications. Bioorg Med Chem 12:2749–2757Google Scholar
  40. 40.
    Clive DL, Hisaindee S, Coltart DM (2003) Derivatized amino acids relevant to native peptide synthesis by chemical ligation and acyl transfer. J Org Chem 68:9247–9254Google Scholar
  41. 41.
    Muir TW, Sondhi D, Cole PA (1998) Expressed protein ligation: a general method for protein engineering. Proc Natl Acad Sci U S A 95:6705–6710Google Scholar
  42. 42.
    Chatterjee C, McGinty RK, Pellois JP, Muir TW (2007) Auxiliary-mediated site-specific peptide ubiquitylation. Angew Chem Int Ed 46:2814–2818Google Scholar
  43. 43.
    McGinty RK, Kim J, Chatterjee C, Roeder RG, Muir TW (2008) Chemically ubiquitylated histone H2B stimulates hDot1L-mediated intranucleosomal methylation. Nature 453:812–816Google Scholar
  44. 44.
    Offer J, Dawson PE (2000) N-alpha-2-Mercaptobenzylamine-assisted chemical ligation. Org Lett 2:23–26Google Scholar
  45. 45.
    Kawakami T, Akaji K, Aimoto S (2001) Peptide bond formation mediated by 4,5-dimethoxy-2-mercaptobenzylamine after periodate oxidation of the N-terminal serine residue. Org Lett 3:1403–1405Google Scholar
  46. 46.
    Vizzavona J, Dick F, Vorherr T (2002) Synthesis and application of an auxiliary group for chemical ligation at the X-gly site. Bioorg Med Chem Lett 12:1963–1965Google Scholar
  47. 47.
    Offer J, Boddy CNC, Dawson PE (2002) Extending synthetic access to proteins with a removable acyl transfer auxiliary. J Am Chem Soc 124:4642–4646Google Scholar
  48. 48.
    Macmillan D, Anderson DW (2004) Rapid synthesis of acyl transfer auxiliaries for cysteine-free native glycopeptide ligation. Org Lett 6:4659–4662Google Scholar
  49. 49.
    Wu B, Chen J, Warren JD, Chen G, Hua Z, Danishefsky SJ (2006) Building complex glycopeptides: development of a cysteine-free native chemical ligation protocol. Angew Chem Int Ed 45:4116–4125Google Scholar
  50. 50.
    Chen J, Chen G, Wu B, Wan Q, Tan Z, Hua Z, Danishefsky SJ (2006) Mature homogeneous erythropoietin-level building blocks by chemical synthesis: the EPO 114–166 glycopeptide domain, presenting the O-linked glycophorin. Tetrahedron Lett 47:8013–8016Google Scholar
  51. 51.
    Kumar KS, Brik A (2010) Accessing posttranslationally modified proteins through thiol positioning. J Pept Sci 16:524–529Google Scholar
  52. 52.
    Brik A, Yang YY, Ficht S, Wong CH (2006) Sugar-assisted glycopeptide ligation. J Am Chem Soc 128:5626–5627Google Scholar
  53. 53.
    Brik A, Wong CH (2007) Sugar-assisted ligation for the synthesis of glycopeptides. Chemistry 13:5670–5675Google Scholar
  54. 54.
    Yan LZ, Dawson PE (2001) Synthesis of peptides and proteins without cysteine residues by native chemical ligation combined with desulfurization. J Am Chem Soc 123:526–533Google Scholar
  55. 55.
    Brik A, Ficht S, Yang YY, Bennett CS, Wong CH (2006) Sugar-assisted ligation of N-linked glycopeptides with broad sequence tolerance at the ligation junction. J Am Chem Soc 128:15026–15033Google Scholar
  56. 56.
    Yang YY, Ficht S, Brik A, Wong CH (2007) Sugar-assisted glycoprotein synthesis. J Am Chem Soc 129:7690–7701Google Scholar
  57. 57.
    Bennett CS, Dean SM, Payne RJ, Ficht S, Brik A, Wong CH (2008) Sugar-assisted glycopeptide ligation with complex oligosaccharides: scope and limitations. J Am Chem Soc 130:11945–11952Google Scholar
  58. 58.
    Pentelute BL, Kent SBH (2007) Selective desulfurization of cysteine in the presence of Cys(Acm) in polypeptides obtained by native chemical ligation. Org Lett 9:687–690Google Scholar
  59. 59.
    Ficht S, Payne RJ, Brik A, Wong CH (2007) Second-generation sugar-assisted ligation: a method for the synthesis of cysteine-containing glycopeptides. Angew Chem Int Ed 46:5975–5979Google Scholar
  60. 60.
    Lutsky MY, Nepomniaschiy N, Brik A (2008) Peptide ligation via side-chain auxiliary. Chem Commun 10:1229–1231Google Scholar
  61. 61.
    Ajish Kumar KS, Harpaz Z, Haj-Yahya M, Brik A (2009) Side-chain assisted ligation in protein synthesis. Bioorg Med Chem Lett 19:3870–3874Google Scholar
  62. 62.
    Payne RJ, Ficht S, Tang S, Brik A, Yang YY, Case DA, Wong CH (2007) Extended sugar-assisted glycopeptide ligations: development, scope and applications. J Am Chem Soc 129(44):13527–13536Google Scholar
  63. 63.
    Payne RJ, Ficht S, Greenberg WA, Wong CH (2008) Cysteine-free peptide and glycopeptide ligation by direct aminolysis. Angew Chem Int Ed 47:4411–4415Google Scholar
  64. 64.
    Thomas GL, Hsieh YSY, Chun CKY, Cai ZL, Reimers JR, Payne RJ (2011) Peptide ligations accelerated by N-terminal aspartate and glutamate residues. Org Lett 13:4770–4773Google Scholar
  65. 65.
    Hojo H, Ozawa C, Katayama H, Ueki A, Nakahara Y, Nakahara Y (2010) The mercaptomethyl group facilitates an efficient one-pot ligation at Xaa-Ser/Thr for (glyco)peptide synthesis. Angew Chem Int Ed 49:5318–5321Google Scholar
  66. 66.
    Chalker JM, Bernardes GJL, Lin YA, Davis BG (2009) Chemical modification of proteins at cysteine: opportunities in chemistry and biology. Chem Asian J 4:630–640Google Scholar
  67. 67.
    Okamoto R, Kajihara Y (2008) Uncovering a latent ligation site for glycopeptide synthesis. Angew Chem Int Ed 47:5402–5406Google Scholar
  68. 68.
    Heinrikson RL (1970) Selective S-methylation of cysteine in proteins and peptides. Biochem Biophys Res Commun 41:967–972Google Scholar
  69. 69.
    Gross E, Morell JL (1974) The reaction of cyanogen bromide with S-methylcysteine: fragmentation of the peptide 14–29 of bovine pancreatic ribonuclease A. Biochem Biophys Res Commun 59:1145–1150Google Scholar
  70. 70.
    Tam JP, Yu Q (1998) Methionine ligation strategy in the biomimetic synthesis of parathyroid hormones. Biopolymers 46:319–327Google Scholar
  71. 71.
    Haase C, Rohde H, Seitz O (2008) Native chemical ligation at valine. Angew Chem Int Ed 47:6807–6810Google Scholar
  72. 72.
    Wan Q, Danishefsky SJ (2007) Free-radical-based, specific desulfurization of cysteine: a powerful advance in the synthesis of polypeptides and glycopolypeptides. Angew Chem Int Ed 46:9248–9252Google Scholar
  73. 73.
    Hoffmann FW, Ess RJ, Simmons TC, Hanzel RS (1956) The desulfurization of mercaptans with trialkyl phosphites. J Am Chem Soc 78:6414Google Scholar
  74. 74.
    Walling C, Basedow OH, Savas ES (1960) Some extensions of the reaction of trivalent phosphorus derivatives with alkoxy and thiyl radicals; a new synthesis of thioesters. J Am Chem Soc 82:2181–2184Google Scholar
  75. 75.
    Walling C, Rabinowitz R (1957) The reaction of thiyl radicals with trialkyl phosphites. J Am Chem Soc 79:5326Google Scholar
  76. 76.
    Rohde H, Seitz O (2010) Ligation-desulfurization: a powerful combination in the synthesis of peptides and glycopeptides. Biopolymers 94:551–559Google Scholar
  77. 77.
    Dawson PE (2011) Native chemical ligation combined with desulfurization and deselenization: a general strategy for chemical protein synthesis. Isr J Chem 51:862–867Google Scholar
  78. 78.
    Chalker JM (2013) Prospects in the total synthesis of protein therapeutics. Chem Biol Drug Des 81:122–135Google Scholar
  79. 79.
    Johnson EC, Malito E, Shen Y, Rich D, Tang WJ, Kent SB (2007) Modular total chemical synthesis of a human immunodeficiency virus type 1 protease. J Am Chem Soc 129:11480–11490Google Scholar
  80. 80.
    Zheng J-S, Tang S, Qi Y-K, Wang Z-P, Liu L (2013) Chemical synthesis of proteins using peptide hydrazides as thioester surrogates. Nat Protocols 8:2483–2495Google Scholar
  81. 81.
    Fang GM, Li YM, Shen F, Huang YC, Li JB, Lin Y, Cui HK, Liu L (2011) Protein chemical synthesis by ligation of peptide hydrazides. Angew Chem Int Ed 50:7645–7649Google Scholar
  82. 82.
    Fang GM, Wang JX, Liu L (2012) Convergent chemical synthesis of proteins by ligation of peptide hydrazides. Angew Chem Int Ed 51:10347–10350Google Scholar
  83. 83.
    Wilkinson BL, Stone RS, Capicciotti CJ, Thaysen-Andersen M, Matthews JM, Packer NH, Ben RN, Payne RJ (2012) Total synthesis of homogeneous antifreeze glycopeptides and glycoproteins. Angew Chem Int Ed 51:3606–3610Google Scholar
  84. 84.
    Garner J, Harding MM (2010) Design and synthesis of antifreeze glycoproteins and mimics. ChemBioChem 11:2489–2498Google Scholar
  85. 85.
    Peltier R, Brimble MA, Wojnar JM, Williams DE, Evans CW, DeVries AL (2010) Synthesis and antifreeze activity of fish antifreeze glycoproteins and their analogs. Chem Sci 1:538–551Google Scholar
  86. 86.
    Sakamoto I, Tezuka K, Fukae K, Ishii K, Taduru K, Maeda M, Ouchi M, Yoshida K, Nambu Y, Igarashi J, Hayashi N, Tsuji T, Kajihara Y (2012) Chemical synthesis of homogeneous human glycosyl-interferon-beta that exhibits potent antitumor activity in vivo. J Am Chem Soc 134:5428–5431Google Scholar
  87. 87.
    Wang P, Dong S, Brailsford JA, Iyer K, Townsend SD, Zhang Q, Hendrickson RC, Shieh J, Moore MAS, Danishefsky SJ (2012) At last: erythropoietin as a single glycoform. Angew Chem Int Ed 51:11576–11584Google Scholar
  88. 88.
    Wang P, Dong S, Shieh J-H, Peguero E, Hendrickson R, Moore MAS, Danishefsky SJ (2013) Erythropoietin derived by chemical synthesis. Science 342:1357–1360Google Scholar
  89. 89.
    Murakami M, Okamoto R, Izumi M, Kajihara Y (2012) Chemical synthesis of an erythropoietin glycoform containing a complex-type disialyloligosaccharide. Angew Chem Int Ed 51:3567–3572Google Scholar
  90. 90.
    Liu S, Pentelute BL, Kent SBH (2012) Convergent chemical synthesis of [lysine24, 38, 83] human erythropoietin. Angew Chem Int Ed 51:993–999Google Scholar
  91. 91.
    Payne RJ (2013) Total synthesis of erythropoietin through the development and exploitation of enabling synthetic technologies. Angew Chem Int Ed 52:505–507Google Scholar
  92. 92.
    Warren JD, Miller JS, Keding SJ, Danishefsky SJ (2004) Toward fully synthetic glycoproteins by ultimately convergent routes: a solution to a long-standing problem. J Am Chem Soc 126:6576–6578Google Scholar
  93. 93.
    Chen G, Warren JD, Chen J, Wu B, Wan Q, Danishefsky SJ (2006) Studies related to the relative thermodynamic stability of C-terminal peptidyl esters of O-hydroxy thiophenol: emergence of a doable strategy for non-cysteine ligation applicable to the chemical synthesis of glycopeptides. J Am Chem Soc 128:7460–7462Google Scholar
  94. 94.
    Wong CTT, Tung CL, Li X (2013) Synthetic cysteine surrogates used in native chemical ligation. Mol Biosyst 9:826–833Google Scholar
  95. 95.
    Botti P, Tchertchian S (2006) Side chain extended ligation. WO/2006/133962Google Scholar
  96. 96.
    Crich D, Banerjee A (2007) Native chemical ligation at phenylalanine. J Am Chem Soc 129:10064–10065Google Scholar
  97. 97.
    Easton CJ, Hutton CA, Tan EW, Tiekink ERT (1990) Synthesis of homochiral hydroxy-α-amino acid derivatives. Tetrahedron Lett 31:7059–7062Google Scholar
  98. 98.
    Easton CJ, Hutton CA, Roselt PD, Tiekink ERT (1994) Stereocontrolled synthesis of β-hydroxyphenylalanine and β-hydroxytyrosine derivatives. Tetrahedron 50:7327–7340Google Scholar
  99. 99.
    Crich D, Banerjee A (2006) Expedient synthesis of threo-beta-hydroxy-alpha-amino acid derivatives: phenylalanine, tyrosine, histidine, and tryptophan. J Org Chem 71:7106–7109Google Scholar
  100. 100.
    Chen J, Wan Q, Yuan Y, Zhu JL, Danishefsky SJ (2008) Native chemical ligation at valine: a contribution to peptide and glycopeptide synthesis. Angew Chem Int Ed 47:8521–8524Google Scholar
  101. 101.
    Chen J, Wang P, Zhu JL, Wan Q, Danishefsky SJ (2010) A program for ligation at threonine sites: application to the controlled total synthesis of glycopeptides. Tetrahedron 66:2277–2283Google Scholar
  102. 102.
    Yang RL, Pasunooti KK, Li FP, Liu XW, Liu CF (2009) Dual native chemical ligation at lysine. J Am Chem Soc 131:13592–13593Google Scholar
  103. 103.
    Marin J, Didierjean C, Aubry A, Casimir JR, Briand JP, Guichard G (2004) Synthesis of enantiopure 4-hydroxypipecolate and 4-hydroxylysine derivatives from a common 4,6-dioxopiperidinecarboxylate precursor. J Org Chem 69:130–141Google Scholar
  104. 104.
    Ajish Kumar KS, Haj-Yahya M, Olschewski D, Lashuel HA, Brik A (2009) Highly efficient and chemoselective peptide ubiquitylation. Angew Chem Int Ed 48:8090–8094Google Scholar
  105. 105.
    El Oualid F, Merkx R, Ekkebus R, Hameed DS, Smit JJ, de Jong A, Hilkmann H, Sixma TK, Ovaa H (2010) Chemical synthesis of ubiquitin, ubiquitin-based probes, and diubiquitin. Angew Chem Int Ed 49:10149–10153Google Scholar
  106. 106.
    Hejjaoui M, Haj-Yahya M, Kumar KS, Brik A, Lashuel HA (2011) Towards elucidation of the role of ubiquitination in the pathogenesis of Parkinson’s disease with semisynthetic ubiquitinated alpha-synuclein. Angew Chem Int Ed 50:405–409Google Scholar
  107. 107.
    Merkx R, de Bruin G, Kruithof A, van den Bergh T, Snip E, Lutz M, El Oualid F, Ovaa H (2013) Scalable synthesis of γ-thiolysine starting from lysine and a side by side comparison with δ-thiolysine in non-enzymatic ubiquitination. Chem Sci 4:4494–4498Google Scholar
  108. 108.
    Kumar KS, Bavikar SN, Spasser L, Moyal T, Ohayon S, Brik A (2011) Total chemical synthesis of a 304 amino acid K48-linked tetraubiquitin protein. Angew Chem Int Ed 50:6137–6141Google Scholar
  109. 109.
    Tan ZP, Shang SY, Danishefsky SJ (2010) Insights into the finer issues of native chemical ligation: an approach to cascade ligations. Angew Chem Int Ed 49:9500–9503Google Scholar
  110. 110.
    Harpaz Z, Siman P, Kumar KSA, Brik A (2010) Protein synthesis assisted by native chemical ligation at leucine. ChemBioChem 11:1232–1235Google Scholar
  111. 111.
    Shang SY, Tan ZP, Dong SW, Danishefsky SJ (2011) An advance in proline ligation. J Am Chem Soc 133:10784–10786Google Scholar
  112. 112.
    Dong S, Shang S, Tan Z, Danishefsky SJ (2011) Toward homogeneous erythropoietin: application of metal free dethiylation in the chemical synthesis of the Ala79-Arg166 glycopeptide domain. Isr J Chem 51:968–976Google Scholar
  113. 113.
    Townsend SD, Tan Z, Dong S, Shang S, Brailsford JA, Danishefsky SJ (2012) Advances in proline ligation. J Am Chem Soc 134:3912–3916Google Scholar
  114. 114.
    Ding H, Shigenaga A, Sato K, Morishita K, Otaka A (2011) Dual kinetically controlled native chemical ligation using a combination of sulfanylproline and sulfanylethylanilide peptide. Org Lett 13:5588–5591Google Scholar
  115. 115.
    Raibaut L, El Mahdi O, Melnyk O (2014) Solid phase protein chemical synthesis. Topics Curr Chem. doi: 10.1007/128_2014_609
  116. 116.
    Siman P, Karthikeyan SV, Brik A (2012) Native chemical ligation at glutamine. Org Lett 14:1520–1523Google Scholar
  117. 117.
    Garner P (1984) Stereocontrolled addition to a penaldic acid equivalent: an asymmetric synthesis of threo-β-hydroxy-L-glutamic acid. Tetrahedron Lett 25:5855–5858Google Scholar
  118. 118.
    Malins LR, Cergol KM, Payne RJ (2013) Peptide ligation-desulfurization chemistry at arginine. ChemBioChem 14:559–563Google Scholar
  119. 119.
    Bang D, Pentelute BL, Kent SB (2006) Kinetically controlled ligation for the convergent chemical synthesis of proteins. Angew Chem Int Ed 45:3985–3988Google Scholar
  120. 120.
    Durek T, Torbeev VY, Kent SB (2007) Convergent chemical synthesis and high-resolution X-ray structure of human lysozyme. Proc Natl Acad Sci U S A 104:4846–4851Google Scholar
  121. 121.
    Torbeev VY, Kent SB (2007) Convergent chemical synthesis and crystal structure of a 203 amino acid “covalent dimer” HIV-1 protease enzyme molecule. Angew Chem Int Ed 46:1667–1670Google Scholar
  122. 122.
    Thompson RE, Chan B, Radom L, Jolliffe KA, Payne RJ (2013) Chemoselective peptide ligation-desulfurization at aspartate. Angew Chem Int Ed 52:9723–9727Google Scholar
  123. 123.
    Guan X, Drake MR, Tan Z (2013) Total synthesis of human galanin-like peptide through an aspartic acid ligation. Org Lett 15:6128–6131Google Scholar
  124. 124.
    Thompson RE, Liu X, Alonso-García N, Pereira PJB, Jolliffe KA, Payne RJ (2014) Trifluoroethanethiol: an additive for efficient one-pot peptide ligation − desulfurization chemistry. J Am Chem Soc 136:8161–8164Google Scholar
  125. 125.
    Rohde H, Schmalisch J, Harpaz Z, Diezmann F, Seitz O (2011) Ascorbate as an alternative to thiol additives in native chemical ligation. ChemBioChem 12:1396–1400Google Scholar
  126. 126.
    Moyal T, Hemantha HP, Siman P, Refua M, Brik A (2013) Highly efficient one-pot ligation and desulfurization. Chem Sci 4:2496–2501Google Scholar
  127. 127.
    Siman P, Blatt O, Moyal T, Danieli T, Lebendiker M, Lashuel HA, Friedler A, Brik A (2011) Chemical synthesis and expression of the HIV-1 Rev protein. ChemBioChem 12:1097–1104Google Scholar
  128. 128.
    Cergol KM, Thompson RE, Malins LR, Turner P, Payne RJ (2014) One-pot peptide ligation-desulfurization at glutamate. Org Lett 16:290–293Google Scholar
  129. 129.
    Malins LR, Cergol KM, Payne RJ (2014) Chemoselective sulfenylation and peptide ligation at tryptophan. Chem Sci 5:260–266Google Scholar
  130. 130.
    Scoffone E, Fontana A, Rocchi R (1966) Selective modification of tryptophan residue in peptides and proteins using sulfenyl halides. Biochem Biophys Res Commun 25:170–174Google Scholar
  131. 131.
    Scoffone E, Fontana A, Rocchi R (1968) Sulfenyl halides as modifying reagents for polypeptides and proteins. I. Modification of tryptophan residues. Biochemistry 7:971–979Google Scholar
  132. 132.
    Wilchek M, Miron T (1972) Conversion of tryptophan to 2-thioltryptophan in peptides and proteins. Biochem Biophys Res Commun 47:1015–1020Google Scholar
  133. 133.
    Shang S, Tan Z, Danishefsky SJ (2011) Application of the logic of cysteine-free native chemical ligation to the synthesis of human parathyroid hormone (hPTH). Proc Natl Acad Sci U S A 108:5986–5989Google Scholar
  134. 134.
    Bock A, Forchhammer K, Heider J, Leinfelder W, Sawers G, Veprek B, Zinoni F (1991) Selenocysteine: the 21st amino acid. Mol Microbiol 5:515–520Google Scholar
  135. 135.
    Lobanov AV, Hatfield DL, Gladyshev VN (2009) Eukaryotic selenoproteins and selenoproteomes. Biochim Biophys Acta 1790:1424–1428Google Scholar
  136. 136.
    Gieselman MD, Xie L, van Der Donk WA (2001) Synthesis of a selenocysteine-containing peptide by native chemical ligation. Org Lett 3:1331–1334Google Scholar
  137. 137.
    Quaderer R, Sewing A, Hilvert D (2001) Selenocysteine-mediated native chemical ligation. Helv Chim Acta 84:1197–1206Google Scholar
  138. 138.
    Hondal RJ, Nilsson BL, Raines RT (2001) Selenocysteine in native chemical ligation and expressed protein ligation. J Am Chem Soc 123:5140–5141Google Scholar
  139. 139.
    Huber R, Criddle RS (1967) Comparison of the chemical properties of selenocysteine and selenocystine with their sulfur analogs. Arch Biochem Biophys 122:164–173Google Scholar
  140. 140.
    Arnold AP, Tan KS, Rabenstein DL (1986) Nuclear magnetic resonance studies of the solution chemistry of metal complexes. 23. Complexation of methylmercury by selenohydryl-containing amino acids and related molecules. Inorg Chem 25:2433–2437Google Scholar
  141. 141.
    Pleasants JC, Guo W, Rabenstein DL (1989) A comparative study of the kinetics of selenol/diselenide and thiol/disulfide exchange reactions. J Am Chem Soc 111:6553–6558Google Scholar
  142. 142.
    Besse D, Siedler F, Diercks T, Kessler H, Moroder L (1997) The redox potential of selenocystine in unconstrained cyclic peptides. Angew Chem Int Ed 36:883–885Google Scholar
  143. 143.
    Nauser T, Dockheer S, Kissner R, Koppenol WH (2006) Catalysis of electron transfer by selenocysteine. Biochemistry 45:6038–6043Google Scholar
  144. 144.
    Metanis N, Keinan E, Dawson PE (2010) Traceless ligation of cysteine peptides using selective deselenization. Angew Chem Int Ed 49:7049–7053Google Scholar
  145. 145.
    Berry SM, Gieselman MD, Nilges MJ, Van der Donk WA, Lu Y (2002) An engineered azurin variant containing a selenocysteine copper ligand. J Am Chem Soc 124:2084–2085Google Scholar
  146. 146.
    Quaderer R, Hilvert D (2002) Selenocysteine-mediated backbone cyclization of unprotected peptides followed by alkylation, oxidative elimination or reduction of the selenol. Chem Commun 2620–2621Google Scholar
  147. 147.
    Malins LR, Mitchell NJ, Payne RJ (2014) Peptide ligation chemistry at selenol amino acids. J Pept Sci 20:64–77Google Scholar
  148. 148.
    Chalker JM, Bernardes GJL, Davis BG (2011) A “tag-and-modify” approach to site-selective protein modification. Acc Chem Res 44:730–741Google Scholar
  149. 149.
    Malins LR, Payne RJ (2012) Synthesis and utility of beta-selenol-phenylalanine for native chemical ligation-deselenization chemistry. Org Lett 14:3142–3145Google Scholar

Copyright information

© Springer International Publishing Switzerland 2014

Authors and Affiliations

  1. 1.School of ChemistryThe University of SydneyCamperdownAustralia

Personalised recommendations