Abstract
Solid phase peptide synthesis (SPPS) provides the possibility to chemically synthesize peptides and proteins. Applying the method on hydrophilic structures is usually without major drawbacks but faces extreme complications when it comes to “difficult sequences.” These includes the vitally important, ubiquitously present and structurally demanding membrane proteins and their functional parts, such as ion channels, G-protein receptors, and other pore-forming structures. Standard synthetic and ligation protocols are not enough for a successful synthesis of these challenging sequences. In this review we highlight, summarize and evaluate the possibilities for synthetic production of “difficult sequences” by SPPS, native chemical ligation (NCL) and follow-up protocols.
Highlights
The “difficult sequence” concept has been introduced in the 80’s and was given distinction by Kent and co-workers for peptides that form strong inter- or intra molecular, non-covalent interactions which form insoluble peptide aggregates
One should consider that the chemical production of “difficult sequences” is composed of several key steps, which include Solid phase peptide synthesis (SPPS), analytical characterization, purification, fragment ligation and if needed post ligation steps (Figure 1)
An impressive example for the value of this strategy was made by Becker and co-workers who used n-octyl glycoside (OG) as an additive for the native chemical ligation (NCL) to synthesize transmembrane peptide diacylglycerol kinase (DAGK) comprising 121 amino acids (Lahiri et al, 2011)
Summary
The “difficult sequence” concept has been introduced in the 80’s and was given distinction by Kent and co-workers for peptides that form strong inter- or intra molecular, non-covalent interactions which form insoluble peptide aggregates. In this review we highlight, summarize and evaluate the possibilities for synthetic production of “difficult sequences” by SPPS, native chemical ligation (NCL) and follow-up protocols. No additional steps are required to remove the solubility tag from the peptide sequence, it is automatically removed at NCL conditions.
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