Synthetic Peptides as Protein Mimics

Synthetic Peptides as Protein Mimics

Synthetic peptides have proven an excellent type of molecule for the mimicry of protein sites. The modified peptides increase the proteolytic stability of the molecules, enhancing their utility for biological applications.

Toolbox for Peptide Synthesis: Non-Proteinogenic Amino Acids and Site-Selective Ligation

The long peptides can be synthesized by the ligation method. Amino acid derivatives with modified backbone length and side-chain orientation, such as d-amino acids, N-alkyl glycine monomers, or proteolytically stable amino acid derivatives can be introduced to the peptides.

Protein Secondary Structure Mimics: α-Helix Mimics, β-Sheet Mimics
Peptide chains can be organized into secondary structures, such as α-helices and β-sheets. Peptides that mimic α-helices and β-sheets of proteins are attractive targets for drug development and tools to explore protein binding mechanism.

The α-helical conformation of a peptide can be induced by adding covalent links between amino acid side chains at selected positions. These links can be formed by lactam and disulfide bridges, triazole-based linkages, and hydrocarbon staples.

In β-sheets, β-strands are connected via loops or turns. Methods to mimic turn structures include macrocyclization, dipeptide of d-proline and l-proline, or α-aminoisobutyric acid in combination with either a d-α-amino acid or an achiral α-amino acid. An example of stimuli-responsive peptides is the temperature-dependent formation of hydrogels by β-sheet peptides. The β-hairpin mimic undergoes gelation upon heating at 60°C, and is completely reversible while cooling.

Protein Mimics in Biomedical Research

Peptides mimicking the CHR region of gp41 were developed to inhibit the formation of the six-helical bundle. Peptides that mimic these receptors are useful tools to explore the details of virus infection mechanism, as well as to develop new drugs against HIV-1. Peptides that mimic the extracellular domains of seven transmembrane G protein-coupled receptors (GPCRs), which is composed of the N-terminus (NT) and the three extracellular loops (ECLs) were explored. Peptide Ac-RERF-NH2 has a high propensity to adopt an α-turn structure and could be a promising drug candidate against cancer.

The design of peptides as protein mimics has evolved as a promising strategy for the exploration of protein-protein interactions, as they are biocompatible, biodegradable, and functionally selective.

Prolong the peptide bioactivity by reversible self-assembly

Self-assembly peptide into fibrillar nanostructures

Most of the potential therapeutic peptides have low solubility, chemical instability or low stability against protease. So it is essential to modify and optimize the peptides to improve the peptide bioavailability.

One novel approach is to create self-assembly, highly ordered and stable nanostructure. For instance, many peptide hormones including glucagon are stored in the form of β-sheet rich amyloid-like fibrils via a hydrogen bond network in the secretory cell.

The oxyntomodulin is a peptide with a potential to treat obesity and diabetes. It is a 37-amino-acid proglucagon-derived peptide hormone with sequence homology to both glucagon and glucagon-like peptide-1 (GLP-1). The oxyntomodulin peptide self-assembles into a stable nanofibril formulation and later on releases active peptide.

Here is the sequence of human oxyntomodulin: His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Met-Asn-Thr-Lys-Arg-Asn-Arg-Asn-Asn-Ile-Ala, HSQGTFTSDY SKYLDSRRAQ DFVQWLMNTK RNRNNIA.

Oxyntomodulin self-assembles into fibrillar nanostructures

  1. The peptide concentration was 10 mg/mL in water at an incubation pH of between 7.0 and 7.3 and low ionic strength (0.09% saline).
  2. The solution was incubated at 37 °C and agitated by orbital shaking.
  3. After incubation for five days, the solution turned turbid due to the formation of a suspension of aggregates. The conversion yield of the self-assembly of Oxyntomodulin into fibrillar nanostructures was estimated to be 99% under these conditions.
  4. The nanofibrils were next used to seed a solution of free peptide at 10 mg/mL in water. 5. The solution was incubated without agitation for one week and then diluted to 1 mg/mL in 0.09% saline for another 2–9 days of incubation at 37 °C and then nine days at room temperature.
  5. Free peptides were mainly in an α-helical conformation. The fibrillar Oxm showed the majority of β-sheet and some α-helical content.

Peptide nanofibrils dissociate to release intact peptide

  1. 1 mg/mL nanofibrils were incubated in water. The 37% of the peptide was released after four-hour incubation.
  2. In aqueous HCl, a 77% release was observed after only four hours. The peptide remained chemically intact after discharge from the nanofibrils

Benefits:

  1. The released peptide is active and nontoxic in vitro.
  2. The nanofibrils prolong peptide serum bioactivity in vivo. And there is no need to engineer or modify the original peptide. Other peptide hormones including glucagon, GLP-1, exendin-4, calcitonin, and gastric inhibitory peptide, are known to self-assemble. The self-assembly method may be used for the clinical application of reversibly self-assembling nanofibrils.

Reference:
Controlling the bioactivity of a peptide hormone in vivo by reversible self-assembly. Nature Communications, volume 8, Article number: 1026 (2017)

Peptide Antigens from Tumor Cells: Potential Peptide Vaccines for Cancer

Check the cancer peptide database for a list of tumor peptides.

Tumor antigens can be classified into two categories based on their pattern of expression: tumor-specific antigens (TSA) and tumor-associated antigens (TAA).

Targeting tumor-associated antigens (TAAs) is a promising approach for cancer immunotherapy. Neoantigens are tumor-specific antigens that originate from somatic mutations in cancer cells but not in healthy tissues. So the TAAs are considered as ideal targets for novel immunotherapies. Antigens of three classes can induce tumor-specific T cell responses.

1. Antigens derived from viral proteins: Viral proteins are produced inside the tumor cells. So the antigenic peptides can be detected by T cells.

2. Antigens derived from point mutations: Many CTL isolated from the tumors were found to recognize antigens that arise from point mutations in ubiquitously expressed genes. These mutations are passenger mutations and the corresponding antigenic peptides are unique to the tumors in which they were identified.

3. Antigens encoded by cancer-germline genes: Cancer-germline genes are expressed in many cancer types and not in normal tissues except germline and trophoblastic cells. The tumor-specific pattern of expression results from the genome-wide demethylation in male germ cells.

A large number of antigenic peptides recognized by antitumor CTL have been identified. Candidate peptides can be synthesized and tested for HLA binding in vitro. The elution of antigenic peptides from MHC class I molecules immunopurified from the surface of tumor cells can be used to identify the antigens. TAAs can be targeted using peptide vaccines or by cellular approaches. The delivery of new peptide drugs might show great promises for future therapies.

Peptide Antigens

Tumor-associated peptide antigens

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How do peptides fold?

peptide fold

Short Peptide Folding

How does the amino acid sequence of a protein chain determine and remain its 3D folded state? How do small proteins fold?

Short Peptide Folding

Many small proteins or miniproteins are peptides shorter than 40-50 residues with stable folding that contain secondary structure elements such as alpha helices and beta strands. An autonomously folding, 35 residue, thermostable subdomain (HP36) of the villin headpiece, is the smallest folded domain of a naturally occurring protein. So, polypeptides simplify the protein-folding problem. It allows in-depth examinations of sequence-structure-stability relationships without using the complex larger proteins. In this recent study, Rocklin et al. designed sequences intended to fold into desired structures. The novel proteins may be useful in bioengineering or pharmacological applications. Check the paper from here: https://goo.gl/Tregb7 http://science.sciencemag.org/content/357/6347/168

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LifeTein Launches Rush Custom Peptide Synthesis Service: Peptide Delivered in 3-5 Days

LifeTein is unveiling an expedited peptide synthesis program, promising to place peptides in its customers’ hands within 3-5 business days. The RushPep™ peptide synthesis service was designed to circumvent the existing limitations of conventional solid-phase peptide synthesis (SPPS), which involves a long coupling time and low yield. RushPep™ shortens the time needed for individual coupling, deprotection and washing steps. The proprietary methodology renders processing ten times faster than in classical synthesis while simultaneously circumventing the limitations caused by the formation of by-products or intermediates to which traditional SPPS approaches are subject.

LifeTein’s Rush Custom Peptide Synthesis Service

“When designing the RushPep™ methodology, our focus was to not only to produce peptides of high quality and purity but also to offer a streamlined solution that would increase the efficiency of researchers’ protein discovery workflows,” stated Dr. Ya Chen, Head of LifeTein’s Rush Peptide Synthesis Group. “RushPep™ achieves these goals by synthesizing the peptides in 3–5 business days to accelerate research and discovery.”

Chen continued, “The reliability of RushPep™ rush peptide synthesis ensures that the peptides are finished in 3–5 business days with high-batch-to-batch reproducibility. ” Most of the crude peptides have a purity of over 80%. RushPep™ peptide service is valuable for the scientists and researchers because it allows them to finish their proteomics projects in a fast and cost-efficient manner.

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Peptides for Parkinson’s disease (PD)

ID2 peptide for inhibition of tumour growth

How does BIRD-2 peptide kill B-cell lymphoma?

Look Who’s Talking

Peptides for Parkinson’s disease (PD)

All peptides used in this study for Parkinson’s disease were synthesized by LifeTein. The α-synuclein is an indication in the pathogenesis of Parkinson’s disease. It was found that a molecular mimicry mechanism between HSV1 and human α-synuclein could trigger PD.

LifeTein Peptides & Parkinson’s Disease

Journal of Neuroimmunology, doi:10.1016/j.jneuroim.2016.01.007 Humoral cross reactivity between α-synuclein and herpes simplex − 1 epitope in Parkinson’s disease, a triggering role in the disease?

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ID2 peptide for inhibition of tumour growth

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LifeTein Launches Rush Custom Peptide Synthesis Service: Peptide Delivered in 3-5 Days

ID2 peptide for inhibition of tumour growth

Biotinylated wild-type and modified (pT27 and T27W) ID2 peptides (amino acids 14–34) were synthesized by LifeTein. ID2 binds to the VHL ubiquitin ligase complex.This ID2 peptide could be used to inhibit tumour growth for patients with glioblastoma.

LifeTein’s ID2 Peptides Can Inhibit Tumour Growth

Nature, 529, 172–177 (14 January 2016) doi:10.1038/nature16475, An ID2-dependent mechanism for VHL inactivation in cancer.

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LifeTein Launches Rush Custom Peptide Synthesis Service: Peptide Delivered in 3-5 Days

LifeTein Peptide Cited in Cell

Peptide library is increasingly used to define antibody epitopes and substrate specificities of protein kinases. For epitope mapping, overlapping peptides are made to span the antigenic protein sequence. The antigenic determinant recognized by a monoclonal antibody can then be screened and defined. The alanine scanning method can also be used to assess that residue’s contribution to antibody binding and to determine which substitutions affect antibody recognition (mutational analysis). Unrelated synthetic peptides can be used to evaluate the antibody cross-reactivity.

Peptide by LifeTein Cited in Cell

Overlapping peptides from LifeTein were used to map the region of Fragment 3 by epitope mapping of anti-Fzd2 antibody. This anti-Fzd2 antibody was found to reduce tumor growth. Wnt signaling plays a critical role in colorectal cancer. Researchers found that Wnt receptor Frizzled2 (Fzd2) and its ligands Wnt5a/b are elevated in metastatic liver, lung, colon, and breast cancer cell lines. Their high level expression correlates with markers of epithelial-mesenchymal transition (EMT). By epitope mapping using synthetic peptides from LifeTein, the researchers mapped the epitope to a specific region. The antibody to Fzd2 was found to reduce cell migration and invasion. Targeting this pathway may provide a cure for patients with tumors expressing high amount of Fzd2 and Wnt5a/b. We have developed an antibody to Fzd2 that reduces cell migration and invasion and inhibits tumor growth and metastasis in xenografts. We propose that targeting this pathway could provide benefit for patients with tumors expressing high levels of Fzd2 and Wnt5a/b.

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Obesity Treatment: A New Peptide Drug

Targeted Drug Melts Fat in Obese Monkeys Currently, only two Food and Drug Administration (FDA) approved drugs for weight loss are available in the United States: the appetite suppressant phentermine and the inhibitor of fat absorption orlistat.

Peptide Drug for Obesity

An MD Anderson group designed a new peptide drug: CKGGRAKDC-GG-D(KLAKLAK)2 (termed adipotide). This is a synthetic peptide that triggers cell death. These data showed that the peptide may be useful for treating obesity in humans. Tthe MD Anderson group used a peptide library to screen and identify regions that bind to specific vascular cells. The interaction identified will be used as effective drugs to target particular protein functions. www.cdc.gov/CDCTV/ObesityEpidemic/ This video explains factors that have contributed to the obesity epidemic.

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Synthetic Peptide Vaccine Research: Problems and Accomplishments

Immune-based cancer treatments are one emerging type of therapy, and they show great potential. Synthetic-peptide-based vaccines, which are designed to elicit T cell immunity, are also a promising approach to the prevention and treatment of both infectious diseases and malignant disorders, such as cancer.

Research on Synthetic Peptide Vaccines

Read more at:http://lifetein.com/therapeutics_peptide_applications.html
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Amino acid composition of cell-penetrating peptides (CPPs)

Obesity Treatment: A New Peptide Drug

Peptide Synthesis for Cell-penetration Studies

D-amino acid peptides to resist common proteases