scholarly journals Amide-forming chemical ligation via O-acyl hydroxamic acids

2018 ◽  
Vol 115 (15) ◽  
pp. 3752-3757 ◽  
Author(s):  
Daniel L. Dunkelmann ◽  
Yuki Hirata ◽  
Kyle A. Totaro ◽  
Daniel T. Cohen ◽  
Chi Zhang ◽  
...  
Keyword(s):  
Peptide Bond ◽  
Amide Bond ◽  
Acyl Transfer ◽  
Medium Size ◽  
Peptide Bonds ◽  
Chemical Ligation ◽  
Native Peptide ◽  
Ligation Product ◽  
Aqueous Conditions ◽  
Acyl Donors

The facile rearrangement of “S-acyl isopeptides” to native peptide bonds via S,N-acyl shift is central to the success of native chemical ligation, the widely used approach for protein total synthesis. Proximity-driven amide bond formation via acyl transfer reactions in other contexts has proven generally less effective. Here, we show that under neutral aqueous conditions, “O-acyl isopeptides” derived from hydroxy-asparagine [aspartic acid-β-hydroxamic acid; Asp(β-HA)] rearrange to form native peptide bonds via an O,N-acyl shift. This process constitutes a rare example of an O,N-acyl shift that proceeds rapidly across a medium-size ring (t1/2 ∼ 15 min), and takes place in water with minimal interference from hydrolysis. In contrast to serine/threonine or tyrosine, which form O-acyl isopeptides only by the use of highly activated acyl donors and appropriate protecting groups in organic solvent, Asp(β-HA) is sufficiently reactive to form O-acyl isopeptides by treatment with an unprotected peptide-αthioester, at low mM concentration, in water. These findings were applied to an acyl transfer-based chemical ligation strategy, in which an unprotected N-terminal Asp(β-HA)-peptide and peptide-αthioester react under aqueous conditions to give a ligation product ultimately linked by a native peptide bond.

Derivatized Amino Acids Relevant to Native Peptide Synthesis by Chemical Ligation and Acyl Transfer

2003 ◽  
Vol 68 (24) ◽  
pp. 9247-9254 ◽  
Author(s):  
Derrick L. J. Clive ◽  
Soleiman Hisaindee ◽  
Don M. Coltart
Keyword(s):  
Amino Acids ◽  
Peptide Synthesis ◽  
Acyl Transfer ◽  
Chemical Ligation ◽  
Native Peptide

scholarly journals Derivatized Amino Acids Relevant to Native Peptide Synthesis by Chemical Ligation and Acyl Transfer.

ChemInform ◽  
2004 ◽  
Vol 35 (13) ◽  
Author(s):  
Derrick L. J. Clive ◽  
Soleiman Hisaindee ◽  
Don M. Coltart
Keyword(s):  
Amino Acids ◽  
Peptide Synthesis ◽  
Acyl Transfer ◽  
Chemical Ligation ◽  
Native Peptide

scholarly journals Peptide Bond Synthesis by a Mechanism Involving an Enzymatic Reaction and a Subsequent Chemical Reaction

2015 ◽  
Vol 291 (4) ◽  
pp. 1735-1750 ◽  
Author(s):  
Tomoko Abe ◽  
Yoshiteru Hashimoto ◽  
Ye Zhuang ◽  
Yin Ge ◽  
Takuto Kumano ◽  
...  
Keyword(s):  
Peptide Synthesis ◽  
Chemical Reaction ◽  
Enzymatic Reaction ◽  
Amide Bond ◽  
Acyl Transfer ◽  
Native Chemical Ligation ◽  
Specific Substrate ◽  
Chemical Ligation ◽  
Subsequent Chemical Reaction ◽  
Subsequent Chemical

We recently reported that an amide bond is unexpectedly formed by an acyl-CoA synthetase (which catalyzes the formation of a carbon-sulfur bond) when a suitable acid and l-cysteine are used as substrates. DltA, which is homologous to the adenylation domain of nonribosomal peptide synthetase, belongs to the same superfamily of adenylate-forming enzymes, which includes many kinds of enzymes, including the acyl-CoA synthetases. Here, we demonstrate that DltA synthesizes not only N-(d-alanyl)-l-cysteine (a dipeptide) but also various oligopeptides. We propose that this enzyme catalyzes peptide synthesis by the following unprecedented mechanism: (i) the formation of S-acyl-l-cysteine as an intermediate via its “enzymatic activity” and (ii) subsequent “chemical” S → N acyl transfer in the intermediate, resulting in peptide formation. Step ii is identical to the corresponding reaction in native chemical ligation, a method of chemical peptide synthesis, whereas step i is not. To the best of our knowledge, our discovery of this peptide synthesis mechanism involving an enzymatic reaction and a subsequent chemical reaction is the first such one to be reported. This new process yields peptides without the use of a thioesterified fragment, which is required in native chemical ligation. Together with these findings, the same mechanism-dependent formation of N-acyl compounds by other members of the above-mentioned superfamily demonstrated that all members most likely form peptide/amide compounds by using this novel mechanism. Each member enzyme acts on a specific substrate; thus, not only the corresponding peptides but also new types of amide compounds can be formed.

scholarly journals Going Round in Circles with N→S Acyl Transfer

Synlett ◽  
2017 ◽  
Vol 28 (13) ◽  
pp. 1517-1529 ◽  
Author(s):  
Derek Macmillan
Keyword(s):  
Transfer Reaction ◽  
Peptide Sequence ◽  
Acyl Transfer ◽  
Native Chemical Ligation ◽  
Full Potential ◽  
Chemical Ligation ◽  
Peptide Engineering ◽  
Naturally Occurring ◽  
Native Peptide ◽  
Peptide Thioesters

It is not highly sophisticated, yet the N→S acyl transfer reaction of a native peptide sequence potentially fills an important technology gap. While several routes to synthetic peptide thioesters exist, only one is routinely applicable for biologically derived samples. Using the naturally occurring amino acid cysteine as the sole activator for N→S acyl transfer we have demonstrated transformation of synthetic and biologically derived precursors into thioesters for use in Native Chemical Ligation, providing a viable alternative for biological samples. Further refinement will be key to realising the full potential of this intriguing process, and increase the number of applications in peptide engineering and therapeutics.1 Introduction2 N→S acyl transfer in ‘normal’ peptide sequences3 Reduced reactivity of internal Xaa-Cys motifs as an advantage in head-to-tail peptide cyclisation4 Reduced reactivity of internal Xaa-Cys motifs as an advantage in modification and cyclisation of biologically produced precursors5 Hydrazinolysis of Xaa-Cys motifs and the acyl hydrazide as a stable thioester equivalent6 Rapid thioester formation via an N→Se acyl shift7 Outlook and conclusions

scholarly journals Isolation, purification and structure of exochelin MS, the extracellular siderophore from Mycobacterium smegmatis

1995 ◽  
Vol 305 (1) ◽  
pp. 187-196 ◽  
Author(s):  
G J Sharman ◽  
D H Williams ◽  
D F Ewing ◽  
C Ratledge
Keyword(s):  
Amino Acids ◽  
Mycobacterium Smegmatis ◽  
Methyl Esters ◽  
Three Dimensional ◽  
Iron Chelation ◽  
Peptide Bond ◽  
Exchange Chromatography ◽  
Peptide Bonds ◽  
Hydrolysis Of ◽  
Gallium Complex

The extracellular siderophore from Mycobacterium smegmatis, exochelin MS, was isolated from iron-deficiently grown cultures and purified to > 98% by a combination of ion-exchange chromatography and h.p.l.c. The material is unextractable into organic solvents, is basic (pI = 9.3-9.5), has a lambda max at 420 nm and a probable Ks for Fe3+ of between 10(25) and 10(30). Its structure has been determined by examination of desferri- and ferri-exochelin and its gallium complex. The methods used were electrospray-m.s. and one- and two-dimensional (NOESY, DQF-COSY and TOCSY) 1H n.m.r. The constituent amino acids were examined by chiral g.l.c analysis of N-trifluoroacetyl isopropyl and N-pentafluoropropionyl methyl esters after hydrolysis, and reductive HI hydrolysis, of the siderophore. The exochelin is a formylated pentapeptide: N-(delta-N-formyl,delta N-hydroxy-R-ornithyl) -beta-alaninyl-delta N-hydroxy-R-ornithinyl-R-allo-threoninyl-delta N-hydroxy-S-ornithine. The linkages involving the three ornithine residues are via their delta N(OH) and alpha-CO groups leaving three free alpha-NH2 groups. Although there are two peptide bonds, these involve the three R (D)-amino acids. Thus the molecule has no conventional peptide bond, and this suggests that it will be resistant to peptidase hydrolysis. The co-ordination centre with Fe3+ is hexadenate in an octahedral structure involving the three hydroxamic acid groups. Molecular modelling shows it to have similar features to other ferric trihydroxamate siderophores whose three-dimensional structures have been established. The molecule is shown to have little flexibility around the iron chelation centre, although the terminal (Orn-3) residue, which is not involved in iron binding except at its delta N atom, has more motional freedom.

Structure of a Novel Enzyme That Catalyzes Acyl Transfer to Alcohols in Aqueous Conditions‡

Biochemistry ◽  
2007 ◽  
Vol 46 (31) ◽  
pp. 8969-8979 ◽  
Author(s):  
Irimpan Mathews ◽  
Michael Soltis ◽  
Mae Saldajeno ◽  
Grant Ganshaw ◽  
Rafael Sala ◽  
...  
Keyword(s):  
Acyl Transfer ◽  
Aqueous Conditions

scholarly journals The complete amino acid sequence of chicken skeletal-muscle enolase

1986 ◽  
Vol 236 (1) ◽  
pp. 115-126 ◽  
Author(s):  
G A Russell ◽  
B Dunbar ◽  
L A Fothergill-Gilmore
Keyword(s):  
Skeletal Muscle ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Peptide Bond ◽  
Peptide Bonds ◽  
Complete Amino Acid Sequence ◽  
Arginine Residues ◽  
Cnbr Cleavage ◽  
Yeast Enolase ◽  
Chicken Skeletal Muscle

The complete amino acid sequence of chicken skeletal-muscle enolase, comprising 433 residues, was determined. The sequence was deduced by automated sequencing of hydroxylamine-cleavage, CNBr-cleavage, o-iodosobenzoic acid-cleavage, clostripain-digest and staphylococcal-proteinase-digest fragments. The presence of several acid-labile peptide bonds and the tenacious aggregation of most CNBr-cleavage fragments meant that a commonly used sequencing strategy involving initial CNBr cleavage was unproductive. Cleavage at the single Asn-Gly peptide bond with hydroxylamine proved to be particularly useful. Comparison of the sequence of chicken enolase with the two yeast enolase isoenzyme sequences shows that the enzyme is strongly conserved, with 60% of the residues identical. The histidine and arginine residues implicated as being important for the activity of yeast enolase are conserved in the chicken enzyme. Secondary-structure predictions are analysed in an accompanying paper [Sawyer, Fothergill-Gilmore & Russell (1986) Biochem. J. 236, 127-130].

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