Serine derived oxazolidines as secondary structure disrupting, solubilizing building blocks in peptide synthesis

Haack, Thomas; Mutter, Manfred

doi:10.1016/s0040-4039(00)91681-2

Cited by 150 publications

(96 citation statements)

References 7 publications

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“…Such modification will probably not change the backbone conformation of a Pro-containing peptide chain. Thus, this pseudo Xaa-Pro bond (27) can generally be viewed as a substituent for any of the Xaa-Pro bonds present in protein sequences and further enlarges the scope of application of our strategy. Of course, it would be also desirable ifthe HMThz derivative could be converted to a natural Cys residue.…”

Section: Methodsmentioning

confidence: 92%

Peptide segment ligation strategy without use of protecting groups.

Liu

Tam

1994

Proc. Natl. Acad. Sci. U.S.A.

210

161

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We describe the concept and the verification of a chemical ligation approach to the synthesis of proteins using peptide se-ents with no protecting groups and no activation of the C-tenal a-carboxyl group. This approach of three steps: (i) aldehyde introduction, in which a masked glycolaldehyde ester Is linked to the carboxyl terminus of an unprotected peptide by reverse protedlysis; (ii) ring formation, in which the umaked aldehyde reacts with the N-teinal a-amino group of the second unprotected peptide containing either a cysteine or a threonine residue to form a t ne or olidine ring at an acidic pH; and (iii) rearrangement in which O-acyl ester linkage is transferred to N-acyl amide linkage to form a peptide bond with a pseudoproline stuture at higher pH. The (4), and the semi-synthesis of proteins consisting of components from synthetic peptides with unusual amino acids and proteins obtained through recombinant technology.The conventional segment-synthesis approaches (5) usually require a maximal protection strategy which has the limitations ofpoor solubility, low coupling efficiency, and the possible danger of racemization due to the overactivation of the carboxyl group. Considerable efforts have been made to overcome these limitations through the use of partial or minimal protecting-group strategies and improvements in the activation methods (6-11). However, these approaches do not overcome the problem of the poor coupling efficiency between large peptide segments, because of the intrinsic difficulty of obtaining effective molar concentrations for high molecular weight molecules. Alternative approaches to overcome the difficulty of forming amide bonds use hydrazone (12,13) or thioether and thioester (14) as surrogate bonds. The advantage of these peptide-bond replacement approaches resides in the high and specific reactivity between the two functional groups not found in amino acids to make the conjugation efficient. The disadvantage is that these linkages are substantially different from the peptide bond, which may lead to conformational distortion, and are usually not stable at either acidic (12) or basic (14) pH.We have now designed and developed a chemical ligation method by which two unprotected peptide segments can be ligated together through an amide bond. In our design, we make use of a highly regiospecific and efficient reaction to link covalently two unprotected peptides. The selectivity of this reaction would also obviate the need for any protecting groups. A peptide bond is then formed in a second step through an intramolecular rearrangement between the two closely neighboring carboxyl and amino groups. The efficiency of the first reaction would solve the slow kinetic problem for reactions between large molecules, and the second reaction, designed as a spontaneous process to form amide bonds without activating the carboxyl group, would overcome the racemization problem of the conventional segment condensation method. EXPERIMENTAL PROCEDURESGeneral. 1H NMR spectra were obtained at 300 ...

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Section: Methodsmentioning

confidence: 92%

Peptide segment ligation strategy without use of protecting groups.

Liu

Tam

1994

Proc. Natl. Acad. Sci. U.S.A.

210

161

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“…These modified amino acid residues were introduced by Mutter and coworkers as temporary protecting groups for peptide synthesis and were found to exert a pronounced effect on the peptide backbone conformation. [1,2] The oxazolidine derivatives are now extensively used as tools to impart better solubility to a growing peptide chain in solid-phase synthesis because they prevent aggregation of growing peptide chains [3,4] and have been found to significantly increase the yields of difficult peptide sequences [5][6][7][8][9] by improving solvation and peptide coupling kinetics. This is attributed to a conformational preference for cis-amide bond formation.…”

Section: Introductionmentioning

confidence: 99%

Solid-State and Solution-Phase Conformations of Pseudoproline-Containing Dipeptides

et al. 2009

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The conformations of 14 threonine-derived pseudoproline-containing dipeptides (including four d-allo-Thr derivatives) have been investigated by NMR. In solution, the major conformer observed for all dipeptides is that in which the amide bond between the pseudoproline and the preceding amino acid is cis. For dipeptides in which the N-terminus is protected, the ratio of cis-to trans-conformers does not depend significantly on the side chain of the N-terminal amino acid, or the stereochemistry of the Thr residue. However, for dipeptides bearing a free N-terminus, there are significant differences in the ratios of cis-to trans-conformers depending on the side chain present. Three dipeptides were crystallized and their X-ray structures determined. In two cases, (benzyloxycarbonyl (Cbz)-Val-Thr(ΨMe,Mepro)-OMe and Cbz-ValThr(ΨMe,Mepro)-OH), the dipeptides adopt a trans-conformation in the solid state, in contrast to the structures observed in solution. In the third case, (9-fluorenylmethoxycarbonyl (Fmoc)-Val-d-alloThr(ΨMe,Mepro)-OH), a cis-amide geometry is observed. These structural differences are attributed to crystal-packing interactions. The conformations of 14 threonine-derived pseudoproline-containing dipeptides (including four d-allo-Thr derivatives) have been investigated by NMR. In solution, the major conformer observed for all dipeptides is that in which the amide bond between the pseudoproline and the preceding amino acid is cis. For dipeptides in which the N-terminus is protected, the ratio of cis-to trans-conformers does not depend significantly on the side chain of the N-terminal amino acid, or the stereochemistry of the Thr residue. However, for dipeptides bearing a free N-terminus, there are significant differences in the ratios of cis-to trans-conformers depending on the side chain present. Three dipeptides were crystallized and their X-ray structures determined. In two cases, (benzyloxycarbonyl (Cbz)-Val-Thr( Me,Me pro)-OMe and Cbz-Val-Thr( Me,Me pro)-OH), the dipeptides adopt a trans-conformation in the solid state, in contrast to the structures observed in solution. In the third case, (9-fluorenylmethoxycarbonyl (Fmoc)-Val-d-allo-Thr( Me,Me pro)-OH), a cis-amide geometry is observed. These structural differences are attributed to crystal-packing interactions.

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“…[6,7] The incorporation into peptide sequences of serine or threonine as oxazolidine, or cysteine as thiazolidine, offers another possibility of disrupting the internal association of the peptide, thus increasing the solvation of the peptide chain. [8][9][10] Our new silicon-containing proline surrogate could be expected to contribute to a better solubility of peptide analogues. In addition, replacing proline with this unnatural amino acid, which is presumably stable towards proteolytic enzymatic degradation, may increase the bioavailability of related peptides without affecting conformational and biological properties considering the structural similarity.…”

Section: Introductionmentioning

confidence: 99%