An improved solid‐phase synthesis of a difficult‐sequence peptide using hexafluoro‐2‐propanol

Milton, Saskia; Milton, Ross D.

doi:10.1111/j.1399-3011.1990.tb00966.x

Cited by 40 publications

(12 citation statements)

References 14 publications

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“…Knowing that the fluoride alcohols are compatible with the SPPS methodology, have the potential to disrupt β‐sheet structure interchains during conventional SPPS39 and have been used in the synthesis of difficult sequences [model peptide H‐Lys 4 ‐Glu 3 ‐Leu 2 ‐Trp(Nps)‐Phe‐OH and the human gonadotropin‐releasing hormone precursor protein40, 41], we then tried to incorporate Gly 124 to the growing peptide‐MBHA by using TBTU/DIPEA in DMSO, 50% TFE/DCM, and 10% HFIP/DCM. The coupling reactions in both fluoride alcohols were performed at room temperature; the reaction carried out in HFIP employed preformed symmetric anhydrides.…”

Section: Resultsmentioning

confidence: 99%

Acanthoscurrin fragment 101–132: Total synthesis at 60°C of a novel difficult sequence

et al. 2008

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Glycine-rich proteins (GRPs) serve a variety of biological functions. Acanthoscurrin is an antimicrobial GRP isolated from hemocytes of the Brazilian spider Acanthoscurria gomesiana. Aiming to contribute to the knowledge of the secondary structure and stepwise solid-phase synthesis of GRPs' glycine-rich domains, we attempted to prepare G(101)GGLGGGRGGGYG(113)GGGGYGGGYG(123) GGY(126)GGGKYK(132)-NH(2), acanthoscurrin C-terminal amidated fragment. Although a theoretical prediction did not indicate high aggregation potential for this peptide, repetitive incomplete aminoacylations were observed after incorporating Tyr(126) to the growing peptide-MBHA resin (Boc chemistry) at 60 degrees C. The problem was not solved by varying the coupling reagents or solvents, adding chaotropic salts to the reaction media or changing the resin/chemistry (Rink amide resin/Fmoc chemistry). Some improvement was made when CLEAR amide resin (Fmoc chemistry) was used, as it allowed for obtaining fragment G(113)-K(132). NIR-FT-Raman spectra collected for samples of the growing peptide-MBHA, -Rink amide resin and -CLEAR amide resin revealed the presence of beta-sheet structures. Only the combination of CLEAR-amide resin, 60 degrees C, Fmoc-(Fmoc-Hmb)Gly-OH and LiCl (the last two used alternately) was able to inhibit the phenomenon, as proven by NIR-FT-Raman analysis of the growing peptide-resin, allowing the total synthesis of desired fragment Gly(101)-K(132). In summary, this work describes a new difficult sequence, contributes to understanding stepwise solid-phase synthesis of this type of peptide and shows that, at least while protected and linked to a resin, this GRP's glycine-rich motif presents an early tendency to assume beta-sheet structures.

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Section: Resultsmentioning

confidence: 99%

Acanthoscurrin fragment 101–132: Total synthesis at 60°C of a novel difficult sequence

et al. 2008

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“…Another promising approach is the addition of 2,2,2trifluoroethanol (TFE) or 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) to N,N-dimethylformamide (DMF) during the coupling steps in order to increase the polarity and solvation properties of the solvent. Examples for this strategy represent the synthesis of d-Ala 17 -phGnRH(14-36) (Milton and Milton, 1990) or model "difficult sequences" (Yamashiro et al, 1976). Fluorinated alcohols are also known as effective solvents for hydrophobic peptides, which can be used during analytics and purification, namely key steps two and three, of the chemical production (Figure 1; Tiburu et al, 2009;Bondarenko et al, 2010;Zhang et al, 2018).…”

Section: External Conditionsmentioning

confidence: 99%

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

Mueller

Baumruck

Zhdanova

et al. 2020

Front. Bioeng. Biotechnol.

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Solid phase peptide synthesis (SPPS) provides the possibility to chemically synthesize peptides and proteins. Applying the method on hydrophilic structures is usually without major drawbacks but faces extreme complications when it comes to "difficult sequences." These includes the vitally important, ubiquitously present and structurally demanding membrane proteins and their functional parts, such as ion channels, G-protein receptors, and other pore-forming structures. Standard synthetic and ligation protocols are not enough for a successful synthesis of these challenging sequences. In this review we highlight, summarize and evaluate the possibilities for synthetic production of "difficult sequences" by SPPS, native chemical ligation (NCL) and follow-up protocols.

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“…Based on previous observations of Narita et al that protected peptide segments in a random coil conformation are more soluble in organic solvent suitable DAEFRHDSGYEVHHQKLVFFAEDVGSNKG AIIGLMVGGVVIA [90] amyloidogenic neurotoxin prion peptide (106-126) KTNMKHMAGAAAAGAVVGGLG [91] acanthoscurrin (101-132) GGGLGGGRGGGYGGGGGYGGGYGGGYG GGKYK-NH 2 [35] Resin for aminoacylation conditions [82,83], Milton et al proposed a method that uses the Chou-Fasman conformational parameter P c (coil parameter for each amino acid) as follows: the average, <P c >, of a peptide segment reveals its tendency to assume a random coil conformation instead of a a-helix or b-sheet structure; consequently, <P c Ã > (P c Ã can be obtained from the linear regression of the function 1/P c ¼ 0.739P a þ 0.345P b ) values greater than 1.0 indicated easy coupling of the subsequent residue in an acceptable time; values in the range of 0.9-1.0 indicated a longer reaction time and need for recoupling; values lower than 0.9 indicated persistent difficult coupling [79]. The use of P a allowed for the preparation of aggregation profiles, such as those of acyl carrier protein (63-74), (Ala) 10 , cytochrome c (66-104), HIV-1 aspartyl protease (86)(87)(88)(89)(90)(91)(92)(93)(94)(95)(96)(97)(98)(99), and growth hormone releasing factor , and for predicting potentially difficult couplings [93]. In 1993, Krchnak et al followed the volume changes of swollen peptide-resins during SPPS and derived a P a for each amino acid coupled (P a : aggregation potential, which reflects the propensity of the amino acid to contribute to peptide aggregation).…”

Section: Difficult Peptide Sequencesmentioning

confidence: 99%

“…Van Woerkon and Van Nispen analyzed the results of 696 couplings using a computer program that considered the Fmoc-amino acid used, its side-chain protecting group, the amino acid to be acylated, the length of the growing peptide, and the result of the Kaiser tests performed [92]. Finally, still in the early 1990s, many reputable researchers recognized that (i) typical difficult sequences present reproducible repetitive incomplete aminoacylations whose improvement by recoupling or capping is limited; (ii) such synthetic difficulty is sequence-dependent, occurring irrespective of resin type or chemical strategy, and aggravated by high resin loadings and sterically hindered amino acids in the sequence; (iii) there is weak or no correlation with the N-terminal amino acid of the growing peptide-resin; and (iv) these symptoms are accompanied by reduced swelling of the growing peptide-resin [78,79,94]. The use of P a allowed for the preparation of aggregation profiles, such as those of acyl carrier protein (63-74), (Ala) 10 , cytochrome c (66-104), HIV-1 aspartyl protease (86)(87)(88)(89)(90)(91)(92)(93)(94)(95)(96)(97)(98)(99), and growth hormone releasing factor , and for predicting potentially difficult couplings [93].…”

Section: Difficult Peptide Sequencesmentioning

confidence: 99%

Difficult Peptides

Miranda

and

Remuzgo

2010

Amino Acids, Peptides and Proteins in Organic Chemistry

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As thousands of biologically active peptides have been discovered in the last 50 years, to date no one contests that all living organismsfrom bacteria to mammals and complex plant formsproduce such compounds for vital purposes. A few examples are given in Table 16.1.In association with the first discoveries of biologically active peptides, it was noted that natural sources could provide quite restricted amounts of them. At that time, the composition and structure of proteins were not sufficiently understood and the ribosomal machinery remained unknown. Hence, many first-rate chemists started searching for methods, procedures, and protocols that could allow for the preparation of peptides in a variable range of scales and purities. Such circumstances turned peptide synthesis into an important research topic [1,2].In the following decades, peptide synthesis reached a high level of efficacy, and also became a fundamental tool for research in the chemistry and biology of these macromolecules [1,3]. Indeed, most of the current knowledge on peptide hormones, antibiotics, cytotoxins, opioids, and hormone-releasing factors has been generated with the help of synthetics.Peptide synthesis has also been critical for providing a wide range of information in biochemistry, medicine, biology, and chemistry as its applications include: (i) therapeutic purposes [4,5]; (ii) development of techniques for peptide detection, separation, identification, quantification, and analysis [6, 7]; (iii) characterization of proteinase-substrate or inhibitor interactions [8,9]; (iv) study of protein-protein interactions and the basis of protein folding [10,11]; (v) mapping of protein epitopes [12, 13]; (vi) mimicking of enzyme activity [14, 15]; (vii) engineering of new peptide-based drugs with therapeutic potential [16, 17]; (viii) design of new biomaterials [18, 19]; (ix) synthesis of prodrugs [20, 21]; and (x) chemical synthesis of proteins [22, 23].Despite these advances, peptide synthesis remains a research topic that attracts an impressive number of scientists worldwide. Most current studies focus

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An improved solid‐phase synthesis of a difficult‐sequence peptide using hexafluoro‐2‐propanol

Cited by 40 publications

References 14 publications

Acanthoscurrin fragment 101–132: Total synthesis at 60°C of a novel difficult sequence

Acanthoscurrin fragment 101–132: Total synthesis at 60°C of a novel difficult sequence

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

Difficult Peptides

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