Synthesis of a highly hydrophobic cyclic decapeptide by solid-phase synthesis of linear peptide and cyclization in solution

A general method was described to synthesize a highly hydrophobic cyclic peptide,cyclo[LWLWLWLWLQ]where underlines indicate D-configuration of the amino acid,by a two-step solid-phase/solution synthesis strategy.The linear decapeptide was assembled by standard Boc chemistry on solid-phase and subsequently cyclized in solution with high efficiency and reproducibility. In subsequent purification by semi-preparative HPLC,50%(v/v) DMF/H_2O was employed as the solvent to overcome the difficulty of solubilization...

Solid-Phase Enzymatic Peptide Synthesis to Produce an Antioxidant Dipeptide

Peptide bond synthesis is favorable to the production of bioactive small peptides. However, the abuse of toxic reagents remains an issue for chemical synthesis method, whereas the low product yield and purity limit the widespread use of enzymatic method. In this study, a new solid-phase enzymatic peptide synthesis(SPEPS) strategy was developed to produce an antioxidant tyrosine-alanine dipeptide(Tyr-Ala) by using recombinant carboxypeptidase Y(CPY) as the catalyst. The general SPEPS procedure involves three steps. First, the N-protected acyl donor was covalently attached to solid resin. Second,the peptide bond was condensed between the acyl donor and the nucleophile under the catalysis of CPY. Finally, one-step cleavage was performed to remove the protecting group and cleave the peptides from solid resin. Upon the optimization of reaction conditions, 77.92%(±2.723%) yield of Tyr-Ala with high product purity of 90.971%(±2.695%) was obtained.In addition, the antioxidant activity of Tyr-Ala was determined by ABTS method, indicating that the synthesized Tyr-Ala obtained by SPEPS showed a superior antioxidant capability compared with commercial glutathione.

Solid-phase Synthesis of PNA Monomer by Ugi Four-component Condensation Reaction

Peptide nucleic acids (PNA) oligomers were synthesized in most cases by peptide synthesis from N-protected monomers. In this work a new method of obtaining PNA monomer by Ugi four-component condensation reaction was tested by solid-phase synthesis. The Fmoc protected PNA monomer was build up with thymin-1-yl acetic acid, 3-methylbutyl aldehyde, Fmoc protected aminoethyl isocyanide and Gly-Wang resin.

Synthesis of Nitrogen-containing Cyclopeptides from N-Terminal Lysine Precursor on Solid Supports

Three synthetic routes for the preparation of nitrogen containing cyclic compounds have been developed, in which the assembling of Fomc-Lys(Boc) residue at N-terminal of resin-bound intermediates is a key prerequisite. Six peptides with nitrogen containing local cyclic structure were efficiently synthesized in good yield starting from chloromethyl resin.

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.

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 of Sea Anemone Peptide Toxin Ap-TxI and Its Insecticidal Activity

[Objectives]This study was conducted to synthesize sea anemone peptide toxin Ap-TxI and investigate its insecticidal activity. [Methods] The sea anemone linear peptide toxin Ap-TxI was synthesized by the solid-phase peptide synthesis(SPPS), and six cysteines were oxidized to form three disulfide bonds by a three-step directional oxidation method. Then, purification by high performance liquid chromatography(HPLC) and mass spectrometry identification were performed. Finally, the insect cytotoxicity and insecticidal activity of Ap-TxI were studied by the MTT method and insect injection method. [Results] The oxidized peptide Ap-TxI with three disulfide bonds in natural configuration was successfully synthesized by the SPPS method, and its purity was >90% by HPLC analysis. The results of the MTT method showed that Ap-TxI was active on the growth of insect cells sf9, with a half effective dose of 0.2 nM;and the results of the mealworm injection test showed that the polypeptide Ap-TxI had high insecticidal activity with a median lethal dose of 11.7 nM. [Conclusions] The sea anemone peptide toxin Ap-TxI with high insecticidal effect was obtained, laying a foundation for the development of new, efficient and safe biological insecticides.

Total synthesis of cyclic heptapeptide euryjanicin B

The first synthesis of the naturally occurring cyclic peptide euryjanicin B has been achieved.A general method was described to synthesize the cyclic peptide by a two-step solid-phase/solution synthesis strategy.All the amino acids in this study are L-configuration. The linear heptapeptide was assembled by standard Fmoc chemistry on solid-phase and subsequently cyclization was carried out by solution method.

Synthesis of two substrate mimics of thioesterase in the biosynthesis of cyclic depsipeptide WS9326A

WS9326 A is a tachykinin receptor antagonist and quorum sensing inhibitor discovered from several Streptomyces strains.The structure of WS9326 A features a(Z)-pentenylcinnamoyl moiety attached on a cyclic depsipeptide skeleton,which is biosynthesized by nonribosomal peptide synthetases(NRPS).The regioselective cyclization in the last step of NRPS catalysis,which is proposed to be catalyzed by a thioesterase(TE)domain in the last module,has not been experimentally characterized.We here report the synthesis of two substrate mimics(1 and 2)of the TE(WS9326 A-TE)in WS9326 A biosynthesis,by using Fmoc-based solid-phase peptide synthesis(SPPS)method.Compounds 1 and 2 are new compounds whose structures have been elucidated based on NMR and HRESIMS analyses.The N-terminal cinnamoyl moiety and C-terminal methylated L-Ser moiety in 2 were incorporated under the mild SPPS conditions.Given the isolation difficulties of substrate of WS9326 A-TE from the Streptomyces producers of WS9326 A,our synthesis of 1 and 2 set the stage for the reconstitution of WS9326 A-TE’s catalytic reaction in vitro in the future.

Use of a Removable Backbone Modification Strategy to Prevent Aspartimide Formation in the Synthesis of Asp Lactam Cyclic Peptides

The synthesis of an Asp lactam derivative of A-183,a selective inhibitor of Factor 7a with good anticoagulant and antithrombotic activity,is described.Our synthesis depends on the use of a removable backbone modification(RBM)strategy to prevent aspartimide formation,which thwarted all attempts to synthesize this target using direct solid-phase peptide synthesis.Validation of the RBM strategy in the synthesis of a second Asp lactam derivative was also accomplished.The RBM strategy is therefore proposed as a general method for the synthesis of Asp lactam cyclic peptides.

STUDIES ON PEPTIDES (Ⅵ)——SYNTHESIS AND PRIMARY BIOLOGICAL ACTIVITY EXAMINING OF THREE FRAGMENTS (93-102, 84-102, 25-38) OF TRANSFORMING GROWTH FACTOR-β

Transforming growth factor-β(TGF-β)is a multifunctional regulatory molecule existing in a wide variety of both normal and neoplastic cells. In order to evaluate the structure-function relationship of TGF-β, we have synthesized a series of its important fragments by manual stepwise solid-phase methodology and studied their biological activity. In

USE OF HOEC ACTIVE ESTERS IN SOLIDPHASE PEPTIDE SYNTHESIS——THE SYNTHESIS OF GROWTH HORMONE RELEASING PEPTIDE

N-hydroxysuccinimide (HOSU) active esters have gained wide application in peptide synthesis, especially in the synthesis of longer peptides by segment conden sation, for they can avoid racemization during coupling processes. However, the HOSU active ester method is prone to side reactions, forming undesirable by-products such as succinimide-oxycarbonyl-alanine N-hydroxysuccinimide ester (Ⅰ) in activating steps and compound (Ⅱ) in the coupling steps sterically hindered by amino acids (proline, valine, isoleucine, etc.).

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）.

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.

An expedient synthesis of peptidyl N-alkylamides by "HOPE" strategy

We have developed an expedient approach,＂HOPE＂（hybrid orthogonal protocol with ease） strategy for the synthesis of peptidyl N-alkylamides.This new strategy was characterized by following points：incorporating Boc and Fmoc protocols together on Merrifield resin,removal of SPG（side-chain protecting groups） without the damage of linker structure on the resin,and the ammonolysis of linker as the last step could achieve the introducing N-alkylamide structure into C-terminal and releasing product from resin-support simultaneously.In present work,eight peptidylamides with different alkylsubstitution at C-terminal were conveniently synthesized by HOPE strategy.

疏水载体辅助液相多肽合成与固相多肽合成工艺的比较

目的分别采用两种液相多肽合成法和固相多肽合成(SPPS)法对具有抗病毒活性的多肽RC-12进行合成工艺研究,通过工艺比较分析以研究其潜在的商业价值。方法分别采用Tag-OH液相法、BDK-OH液相法及SPPS法合成抗病毒多肽RC-12,并对三种不同的合成工艺进行物料消耗量、“三废”产生量、收率比较。结果比较Tag-OH液相法和BDK-OH液相法,两种方法的收率相差不大且都优于SPPS法;BDK-OH液相法物料消耗量和“三废”产生量高于Tag-OH液相法,但都远远低于SPPS法。结论Tag-OH液相法对合成多肽RC-12具有较好的工业研究意义和潜在的商业价值。

生物活性肽研究进展

氨基酸彼此以酰胺键相连而形成的化合物称作肽,并有寡肽和多肽之分,前者一般含10个以下的氨基酸残基,后者含10-50个氨基酸残基。由于肽的相对分子量对于蛋白质小而且表现为一定的生物活性如降血压、提高免疫、抗癌等,故而称为生物活性肽。伴随分子生物学、生物化学技术的飞速发展,人们对生物活性肽的认识和利用水平得到不断的提高。文中从生物活性肽的来源、制备、功能及应用等方面,对生物活性肽的研究作了一个比较全面的综述。

构象限制二肽的合成及其晶体结构研究

以L-苯丙氨酸甲酯和消旋的1-(4-甲氧苯基)-2,5-二苯基-6-氧-4-氮螺[2.4]-4-烯-7-庚酮为原料,采用噁唑酮法合成了两个含2,3-二芳基取代环丙烷氨基酸残基的构象限制二肽的非对映异构体,并用单晶X-射线衍射法对这两个二肽非对映体的结构进行研究。

混合酸酐法合成CCK-4

目的液相合成Trp-Met-Asp-PheNH2四肽(CCK-4)。方法以N-保护的色氨酸、甲硫氨酸、天门冬氨酸和C-保护的苯丙氨酸为原料,采用混合酸酐法合成了CCK四肽,总收率38.5%。结果其结构经IR、1HNMR、MS确证,熔点与文献报道的一致。结论采用重复过量混合酸酐法合成了目标化合物(6),所用方法经济、有效、可靠。