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.
Synthesis of multiple antigenic peptides: strategies and limitations
Dendrimeric platforms such as multiple antigenic peptides (MAP) can be synthesized either entirely by solid-phase methods (SPPS, direct approach) or by conjugation in SPPS-made building blocks (indirect approach).
Synthesis of Multiple Antigenic Peptides
MAP peptide synthesis
http://www.ncbi.nlm.nih.gov/pubmed/21391284
SPPS is the preferred method by LifeTein. The synthesis approach requires a branched poly-lysine core. Each branch is elongated into the corresponding epitope by stepwise SPPS. The disadvantage of this approach is that the synthetic errors could happened and cause microheterogeneity in the final materials. However the cost is lower and less time-consuming than the indirect approach. For very long linear peptides, it is more advantageous to use the SPPS method.
The MAP synthesis may not always meet with success. The solubility of the peptide epitope can also become an issue and is difficult to predict for long epitopes. It is recommended to carefully design and analyze the linear epitope before MAP synthesis.
Studies showed that synthesis with Ahx linker in the lysine core had better isolated yield. It is possible that the flexibilizing effect of Ahx helps in keeping peptide chains properly solvated during synthesis, preventing aggregation and hence increasing the amount of viable growing peptide sequences.
Peptide protein binding strategies: Structure, Volume 18, Issue 2, 188-199, 10 February 2010
Peptide-Protein Binding
Highlights
After binding its partners, most peptides do not introduce any conformational changes
The interfaces of peptide-protein have more hydrogen bonds
The peptide hotspots are important for the binding
Peptides prefer to bind in the largest pockets on the protein surface
LifeTein is pleased to offer a free, comprehensive web-based peptide analysis tool. This tool will allow your research team to overcome common difficulties inherent in protein analysis and peptide antigen design.
Proteins and most naturally occurring peptides are composed of amino acids in the L-configuration. However, D-amino acids have been detected in a variety of peptides synthesized in animal cells. Examples include opiate and antimicrobial peptides from frog skin, neuropeptides from snails, hormones from crustaceans, and venom from spiders. These D amino acid peptides are considered to be the most promising alternative for anticancer, anti-inflammatory, antimicrobial, and delivery agents.
About D-Amino Acid Peptides
The design of all-D-peptides has been applied to increase bioactive peptides’ resistance to endogenous enzymes, as well as their bioavailability. Retro-inverso peptides are obtained by replacing the standard L-amino acid residues with the corresponding D-amino acids and reversing the direction of the peptide backbone. Therefore, the original spatial orientation and the chirality of the side chains is unchanged. This results in a non-complementary side chain topochemistry between the analog and the parental L-peptide. The significantly improved biostability of D-peptides usually leads to longer in vivo circulation half-time, making the D-peptide based drug delivery system more attractive and efficient than their L-peptide counterparts.
Success has been achieved immunologically in using retro-inverso peptides toward antigenic mimicry of their parent l-peptides. It was found that the retro-all-d-peptide isomer of p53(15–29), like its parent l-peptide, adopted a right-handed helical conformation in the complex. However, in some cases, the retro-inverso isomers are significantly inferior to their parent l-peptides. The low cellular uptake of D-peptides remain an unmet challenge.
So the best way is to insert some key D amino acids into the peptide sequence. For example, some pharmaceutical important peptide antibiotics such as gramicidins, actinomycins, or bacitracins incorporate D-amino acids into the drug design. The assemblies of D-peptides deserve further exploration and may lead to more surprises.
D amino acid peptide with high stability
See more details from here: http://lifetein.com/Peptide-Synthesis-D-Amino-Acid.html
Reference: http://www.pnas.org/content/102/2/413.full.pdf+html
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Cell-penetrating peptides (CPPs) such as the HIV TAT peptides are able to enter cells by direct translocation and endocytosis. Click here to see details about the CPP: http://lifetein.com/Cell_Penetrating_Peptides.html
About Cell-Penetrating Peptides
Cell Penetrating Peptides
The following table shows a selection of currently known CPPs, their origins, and sequences.
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 (adipotide). This is a synthetic peptide that triggers cell death. These data showed that the peptide might be helpful in treating obesity in humans. The 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. This video explains factors that have contributed to the obesity epidemic.
Cell-penetrating peptides (CPPs) have the ability to enter a cell’s plasma membrane independent of a membrane receptor. Attached to a CPP, therapeutic cargo could be delivered to an intracellular target, thus overcoming the entry restrictions set by the plasma membrane.
Peptide Synthesis & Cell Penetration
The cationic CPPs interact with negatively charged head groups of lipids directly in the plasma membrane through electrostatic interactions. The increased local peptide concentration at the membrane surface will cause a transient destabilization of the lipid bilayer and lead to cell entry. The hydrophobic interactions, especially facilitated by the presence of tryptophan residues, may be important for the CPP-membrane interaction and cellular internalization.
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