A NOVEL COUPLING REAGENT(PENTAFLUOROPHENYL DIPHENYLPHOSPHPATE)FOR THE PEPTIDE SYNTHESIS

Pentafluorophenyl diphenylphosphate was found to be a novel convenient coupling reagent for the peptide synthesis with several advantages.Higher yield and lower racemization were found with this reagent than DCCI.

Application of an Organophosphorus Compound-DEPBT as Coupling Reagent in Liquid Phase Peptide Synthesis

DEPBT, a novel coupling reagent containing phosphorus, was used in synthesis of pentapeptide Boc-Ile-Asn-Met-Trp(For)-Gly.OMe by solution method.

An Unusual Acceleration of Amination Reactivity by the Proximal Ester Groups in Catechol Diesters: An Efficient Way for Peptide Synthesis

Peptide bond formation has drawn more and more attention due to its importance in medicinal chemistry and drug discovery.Using active esters to construct peptide bonds is an indispensable strategy in this field.We herein report a fast and efficient amination reaction to synthesize peptides from catechol diesters.Catechol monoesters are good active esters in peptide synthesis due to the anchimeric assistance effect.In previously reported work,the amination reaction was inactivated if the 2-hydroxyl group in the catechol esters was functionalized.However,we found an unusual acceleration of the amination reactivity when the 2-hydroxyl group was functionalized as an ester group.The reaction can be complete in 2−3 min in some cases.This acceleration was found through experimental studies to not result from the anchimeric assistance effect.We proposed aπ*−π*interaction pathway between the two proximal ester groups.Some computational studies were made to support this assumption.

Study on Chemical Synthesis and Insecticidal Activity of Peptideκ-CTx-btg02

[Objectives]This study was conducted to investigate the chemical synthesis of peptideκ-CTx-btg02.[Methods]Linear peptideκ-CTx-btg02 was synthesized by solid phase peptide synthesis(SPPS).After oxidation and folding of the linear peptide,mass spectrometry identification and high performance liquid chromatography(HPLC)purification were performed.Then,the MTT method and insect injection method were applied to study its insecticidal activity.[Results]The peptideκ-CTx-btg02 was successfully synthesized by the SPPS method,and identified for the formation of disulfide bonds by mass spectrometry identification,and the purity after HPLC separation was greater than 95%.The MTT experiment showed that the peptideκ-CTx-btg02 could inhibit the growth of insect cells sf9,with a half effective dose of 0.13 nM.The insect injection experiment showed that the peptideκ-CTx-btg02 could effectively kill Tenebrio molitor,with a half lethal dose of 15.6 nM.The results of the electrophysiological experiment showed that 10μM peptideκ-CTx-btg02 had no blocking activity on murine acetylcholineα2β3 andα3β4.Therefore,the peptideκ-CTx-btg02 had a good inhibitory effect on the growth of insect cells,highly effective insecticidal activity and weak mammalian toxicity.[Conclusions]This study lays a foundation for the development of new,safe and efficient peptide biological insecticides.

Chemo-enzymatic Synthesis of Z-Asp-Val-Tyr-NH_2——A Precursor Tripeptide of Thymopentin

Z-Asp-Val-Tyr-NH2,a precursor tripeptide of thymopentin（TP-5）,was successfully synthesized by a combination of chemical and enzyme methods.Firstly,the precursor dipeptide Val-Tyr-NH2 was synthesized by a novel chemical method in four steps including the preparation of NCA-Val,the esterification of L-tyrosine and the ammonolysis of L-tyrosine ethyl ester,and the formation of Val-Tyr-NH2 from NCA-Val and L-Tyr-NH2.Secondly,a thermolysin,thermoase PC 10F was used to catalyze the formation of Z-Asp-Val-Tyr-NH2 in an aqueous/organic solvent diphase system with Z-Asp-OH and Val-Ty-NH2 as the substrates under thermodynamic control.The optimum conditions were pH=6.5,40 ℃,thermoase 100 mg,in 2-（N-morpholino） ethanesulfonic acid MES/NaOH（containing 5 mmol/L CaCl2） of n-butanol system（15/85,volume ratio） for 20 h in a maximum yield of 27.02%.

Peptide synthesis using pentafluorophenyl diphenylphosphate (FDP) as coupling reagent

Comparison of the reaction speed and yield with its analogues and some conventional peptide coupling reagents, pentafluorophenyl diphenylphosphate (FDP) was shown to be a more preferable ''active ester'' type reagent for the peptide synthesis. The synthesis of oligopeptides using FDP was achieved with high yields. The influences of several reaction parameters such as solvent, base, additive and temperature on the coupling reaction were studied using HPLC method. The degree of racemization with FDP determined by HPLC or Young test was shown to be lower than that of DCCI. Octapeptide Gly-Cys(Bzl)-Ser-Gly-Lys-Leu-Ile-Cys(Bzl)-OH, corresponding to the amino acid sequence of gp41 of HIV-1, was successfully synthesized by 5+3 approach using FDP with high yield.

SYNTHESIS OF CYCLIC PEPTIDES USING BBC REAGENT

We report here the synthesis of cyclic peptides using benzotriazolyloxy-bis(pyrrolidino)-carbonium hexafluoro- phosphate(BBC)as a coupling reagent with high yield and speed even in low concentration compared with the other usual coupling reagents.

SYNTHESIS OF PEG-DENDRIMERIC PEPTIDE

Synthesis of a new macromolecule with MW up to 7.6 KDA is described in the present paper. The ligation between a dendrimeric amino-PEG and unprotected peptides via a site- specific reaction is a useful trial for the preparation of synthetic vaccine and related compounds.

De Novo Design and Synthesis of Four-Helix Bundle Peptides

ＤｅＮｏｖｏＤｅｓｉｇｎａｎｄＳｙｎｔｈｅｓｉｓｏｆＦｏｕｒ－ＨｅｌｉｘＢｕｎｄｌｅＰｅｐｔｉｄｅｓＤｅＮｏｖｏＤｅｓｉｇｎａｎｄＳｙｎｔｈｅｓｉｓｏｆＦｏｕｒ－ＨｅｌｉｘＢｕｎｄｌｅＰｅｐｔｉｄｅｓ￥ＸｉａｎｇＱｕｎＬＩ；ＺｈｅｎＷｅｉＭＩＡＯ；Ｙ...

SYNTHESIS OF NEW ESTRONE DERIVATIVES OF AMINO ACIDS AND PEPTIDES

A series of estrone derivatives of amino acids and peptides haze been synthesized by different coupling reagents and the binding affinity of deblocked derivatives to the estrogen receptor of rats uteri have been measured by a comptetive radiometric bind- ing assay.

Protease-catalyzed Synthesis of Bz-Arg-Gly-Asp-OMe in Full Aqueous Medium

Synthesis of N-benzoyl-argininylglycylasparagine methyl ester（ Bz-Arg-Gly-Asp-OMe）, a precursor tripeptide of Arg-Gly-Asp） was catalyzed by papain under kinetic control, at alkaline pH, in a full aqueous medium. The substrates were N-benzoyl-argininylglycine ethyl ester and asparagine dimethyl ester. An aqueous solution of 0. 1 mol/L KCl/NaOH containing 8 mmol/L EDTA and 2 mmol/L DTT was selected as the reaction medium. The synthesized hydrophilic tripeptide was soluble in the reaction medium during the reaction process, however, the secondary hydro- lysis of the tripeptide product was not considerable. The effects of different factors, including water content, temperature, reaction time, and molar ratio of the substrates, on the yield of Bz-Arg-Gly-Asp-OMe were examined. The optimal reaction conditions were 0. 05 mol/L Bz-Arg-Gly-OEt and 0. 15 mol/L Asp（ -OMe）2·HCl in 0. 1 mol/L KCl/ NaOH solution（pH 8.5）, at 40 ℃, and a reaction time of 60 min, with a maximum conversion yield of 62.4%.

An Improved Synthesis of Laminin Fragment CR9

Laminin nonapeptide CR9 was synthesized via two different methods. A notably enhanced yield (46.8%) was obtained from method B compared to that (12.4%) from standard protocol (method A).

Preparation and Insecticidal Activity of Sea Anemone Peptide AP-GI from Aiptasia pallida

[Objectives] This study was conducted to synthesize sea anemone peptide Ap-GI and investigate its insecticidal activity. [Methods] The linear peptide Ap-GI was synthesized by solid phase peptide synthesis(SPPS), and the linear peptide was subjected to the two-step oxidative folding, mass spectrometry identification and high performance liquid chromatography(HPLC) purification. Then, the MTT method and insect injection method were used to study its insecticidal activity. [Results] The synthesized sea anemone peptide had a purity of 95%. The test results of the MTT method showed that the peptide Ap-GI had the activity of inhibiting the growth of insect cells sf9 with the median effective dose of 0.7 nM;and the test results of the injection method on yellow mealworms showed that the peptide Ap-GI had high insecticidal activity, and the median lethal dose was 16.9 nM. [Conclusions] The sea anemone peptide Ap-GI from Aiptasia pallida has a good inhibitory effect on the growth of insect cells and high-efficiency insecticidal activity, which can lay a foundation for the development of new, safe and efficient peptide biological insecticides.

Recent advances in chemical protein synthesis:method developments and biological applications

The central dogma of modern biology underscores the pivotal roles proteins play in diverse biological processes,the study of which necessitates advanced methods to produce proteins with precision and versatility.Chemical protein synthesis,a powerful approach utilizing chemical reactions for the de novo construction of structurally accurate proteins,has emerged as a transformative tool for studying proteins and generating protein derivatives/mimics inaccessible by natural biological machinery,including post-translationally modified proteins,proteins comprised of unnatural amino acids,as well as mirror-image proteins.This review summarizes recent strides in synthetic method developments for chemical protein synthesis,including innovative techniques in solid-phase peptide synthesis,the challenges presented by difficult sequences in either synthesis or folding and the exploration of novel ligation reactions using both chemical and enzymatic methods.Furthermore,the review also delves into newly developed protocols for site-selective protein modifications and the generation of stapled or macrocyclized peptides/miniproteins,highlighting the power of chemical methods to make structurally diverse proteins.Recent applications of synthetic proteins in investigating post-translational modifications(phosphorylation,lipidation,glycosylation,ubiquitination,etc.),mirror-image biological processes and drug development are further discussed.Together,these topics provide a comprehensive overview of the current landscape of chemical protein synthesis.

An improved installation of 2-hydroxy-4-methoxybenzyl(iHmb)method for chemical protein synthesis

The 2-hydroxy-4-methoxybenzyl(Hmb)backbone modification can prevent amide bond-mediated sidereactions(e.g.,aspartimide formation,peptide aggregation)by installing the removable Hmb group into a peptide bond,thus improving the synthesis of long and challenging peptides and proteins.However,its use is largely precluded by the limited Hmb’s installation sites.In this report,an improved installation of Hmb(iHmb)method was developed to achieve the flexible installation and the convenient removal of Hmb.The iHmb method involves two critical steps:(1)oxidative diazotization of the readily installed 2-hydroxy-4-methoxy-5-amino-benzyl(Hmab)to give 2-hydroxy-4-methoxy-5-diazonium-benzyl(Hmdab)by combining soamyl nitrite(IAN)/HBF_(4),and(2)reductive elimination of Hmdab to give the desired Hmb by 1,2-ethanedithiol(EDT).The iHmb method enables the installation of Hmb at any primary amino acid including the highly sterically hindered amino acids(e.g.,valine and isoleucine).The practicality and utility of the iHmb method was demonstrated by one-shot solid-phase synthesis of a challenging aspartimide-prone peptide,the mirror-image version of a hydrophobic peptide and a long-chain peptide up to 76-residue.Furthermore,the iHmb method can be utilized to facilitate chemical protein ligation,as exemplified by the synthesis of the single-spanning membrane protein sarcolipin.The iHmb method expands the toolkit for peptide synthesis and ligation and facilitates the preparation of peptides/proteins.

First Total Synthesis and Biological Potential of a Heptacyclopeptide of Plant Origin

Synthesis of a natural glycine-rich heptacyclopeptide mahafacyclin A （7） was accomplished by solution-phase technique of peptide synthesis via coupling of tetrapeptide unit Boc-L-Thr-L-Ile-L-Leu-Gly-OH with tripeptide unit L-Val-L-Phe-Gly-OMe followed by cyclization of linear heptapeptide fragment. Structure of the newly synthesized cyclopolypeptide was confirmed by means of chemical, spectroscopic analyses and subjected to antibacterial, antifungal and anthelmintic activity studies. Bioactivity results showed potent antifungal and anthelmintic activities of the synthesized peptide against dermatophytes T. mentagrophytes, M. audouinii and earthworm species M. kon-kanensis, P. corethruses and E. eugeniea.

Novel Chemical Strategy for the Synthesis of RGDCySS Tetrapeptide

A new route was described to synthesize Arg-Gly-Asp-X（RGDX,X=amino acid） tetrapeptide.To better understand the method,the tetrapeptide Arg-Gly-Asp-CySS（RGDCySS） was chosen as a model target for X.First,GDCySS was obtained in four steps,comprising the chloroacetylation of L-aspartic acid（ClCH2COAsp）,synthesis of chloroacetyl L-aspartic acid anhydride[ClCH2COAsp（CO）2O],formation of ClCH2COAsp-CySS and ammonolysis of ClCH2COAsp-CySS.Second,preparation of Arg-NCA,which was coupled with GDCySS to synthesize RGDCySS by the NCA method（Leuchs＇ anhydrides method,NCA：N-carboxy-a-amino acid anhydride）.The purity of the product was analyzed by the high performance liquid chromatography（HPLC）.Molecular weights of the peptide products were confirmed by mass spectroscopy.In the developed approach,less protected amino acids were used compared to conventional solid-phase synthesis.The new route offers advantages of low cost,simplicity and rapid synthesis with a reasonable yield of 63.0％（calculated according to arginine content）.

Enzymatic Synthesis of a CCK-8 Tripeptide Fragment

A process for the synthesis of CCK-8 tripeptide H-Gly-Trp-Met-OH catalyzed by immobilized enzyme was re-ported. Enzymes were used for the formation of peptide bonds and the removal of protecting group. Starting with phenylacetyl (PhAc) glycin, N-protected dipeptide PhAc-Gly-Trp-OMe was obtained by coupling PhAc-protected glycine carboxamidomethyl ester (OCam) with Trp-OMe catalyzed by immobilized papain in buffered ethyl acetate. Then the condensation between PhAc-Gly-Trp-OMe and Met-OEtHCl was carried out by immobilized -chy-motrypsin catalysis in solvent free system. Basic hydrolysis was followed getting PhAc-Gly-Trp-Met-OH. The PhAc-group was removed with penicillin G amidase and H-Gly-Trp-Met-OH was obtained in an overall yield of 43.9%. The reaction conversion of tripeptide in solvent free system was strongly affected by the system of basic salts added. The influence of the support materials used to deposit enzymes and structures of acyl donor and nu-cleophile on the reaction was also investigated.

Exploiting Complex-Type N-Glycan to Improve the in Vivo Stability of Bioactive Peptides

Peptides can be potentmolecules with high efficacy and selectivity in the development of biotherapeutics.However,the poor pharmacokinetic properties of peptides pose major challenges for their broader medicinal applications.Inspired by the proteinstabilizing role of natural N-glycosylation,we design and synthesize a series of parathyroid hormone(PTH)peptides(1-34),bearing either N-GlcNAc or biantennary complex-type N-glycan modification,and evaluate their serum stability and biological activities.The results indicate that an N-Asn-linked complex-type sialylundecasaccharide can increase the serum half-life and in vivo bioactivity of PTH peptides with a broad tolerance of modification sites.Further,hydrogen/deuterium exchange mass spectroscopy indicates that the larger-sized Nglycan can induce enhanced hydration dynamics in its surroundings,which may facilitate an improved resistance for the peptide against enzymatic proteolysis.This sialylundecasaccharide-based peptideengineering strategy has also been applied to glucagon-like peptide-1(7-37),leading to glycopeptides with enhanced hypoglycemic activity and acting time in vivo.Together,these results demonstrate the potential of using sialylated complextype N-glycan as a general engineering strategy for developing long-acting peptide therapeutics.

Solid phase synthesis of peptide–siRNA conjugates containing disulfde bond unit

A disulfide-modified nucleoside was designed and synthesized. After loading the modified nucleoside on controlled pore glass （CPG）, solid phase synthesis strategy was used to prepare peptide-oligonucleotide conjugates （N-3＇） containing disulfide bond unit. The 3＇-sense strand peptide-siRNA conjugate （VII） maintained good gene silencing activity, while that of the 3＇-antisense strand conjugate decreased somewhat. And the sense strand off-target effect of VII decreased remarkably.