Key Points Of Peptide Design

Basic Principles of Peptide Design

Design Solutions for Peptides That are Difficult to Synthesize

When designing peptides, factors such as peptide length, solubility, secondary structure, and peptide composition need to be considered.

• Peptide length. The length of peptides is variable and depends on where they are used. For example, peptides with 10-20 amino acid residues are suitable for antibody preparation, while peptides used for structure/function research are more flexible in length. However, the longer the peptide sequence, the more difficult it is to synthesize the peptide, and the purity of peptide synthesis will decrease as the length of the peptide sequence increases. Generally speaking, a peptide length of 15-20 amino acids is moderate, and peptides with a length of less than 30 amino acids can basically be synthesized normally. If the peptide exceeds 40 amino acids, the success rate of synthesis will drop a lot. For peptides that are difficult to synthesize, our professionals will give customers reasonable suggestions after careful analysis and research.

• Solubility. Peptide solubility is an important aspect of peptide design. Peptides containing a high proportion of hydrophobic residues, such as leucine, valine, isoleucine, methionine, phenylalanine and tryptophan, have limited or no solubility in aqueous solutions. These peptides are then difficult to use in experiments and are difficult to purify. We recommend keeping the proportion of hydrophobic amino acids in the peptide below 50% and that at least one out of every five residues be charged.

• Secondary structure. When designing peptides, we must consider the formation of β-folds. During the synthesis process, as the peptide chain is extended, the β-fold structure leads to a lot of missing sequences in the final product. We recommend choosing sequences that do not contain a large number of consecutive valine, isoleucine, tyrosine, phenylalanine, tryptophan, leucine, glutamine, and threonine to avoid such problems. If sequences containing these residues cannot be avoided, we can insert a glycine or proline every 3 residues, replace glutamine with asparagine, or replace threonine with serine.  

• Peptide composition. Certain amino acids or amino acid combinations can also have a negative impact on peptide synthesis and purification. It is difficult to obtain high purity peptides containing multiple cysteines, methionines, or tryptophans because the side chains of these amino acids are easily oxidized or undergo side reactions. If possible, try to minimize the presence of these residues in the peptide. Usually, we can use norleucine instead of methionine and serine instead of cysteine.

• Shorten the sequence. Generally speaking, the longer the peptide chain, the lower the purity of the crude peptide. In most cases, shortening the peptide to less than 30 residues can achieve the desired results.
• Reduce the number of hydrophobic residues. If the sequence contains a large number of hydrophobic residues, especially when they are distributed at positions 7-12 from the carboxyl end, the peptide will be difficult to synthesize. This may be caused by incomplete coupling caused by the formation of β-folds by the side chains of the peptide during the synthesis process. In this case, replacing one or more hydrophobic residues with some polar residues, or inserting a glycine or proline, can open the β-fold.
• Minimize "difficult" residues. If the sequence contains a large number of cysteine, methionine, tryptophan residues, the peptide will be difficult to synthesize. Because cysteine, methionine, tryptophan or their side chains are easily oxidized. If possible, try to avoid these residues in the sequence, or make some conservative substitutions.