Solid-phase synthesis of sterically hindered peptides via ribosome-mimicking molecular reactors
Peptides containing sterically hindered amino acids are attractive drug leads due to their enhanced stability and bioavailability. Here, a protocol is described to efficiently synthesize such peptides by mimicking riboso
The incorporation of N-methylated or α,α-disubstituted amino acids into peptides can markedly enhance their drug-like properties by improving proteolytic stability and membrane permeability. However, the solid-phase synthesis of such sterically hindered peptides remains a persistent challenge. This paper presents a detailed protocol based on a recently developed ribosome-mimicking molecular reactor strategy, which substantially improves the synthetic efficiency of these difficult sequences via solid phase peptide synthesis. The protocol provides a comprehensive, step-by-step procedure to facilitate a quick startup by users, including the synthesis of Oxyma-C as the key activating unit precursor in the RMMR structure. The preparation of RMMR resin is achieved through modification of conventional SPPS resin, and the implementation of RMMR-SPPS is demonstrated in both manual and automated synthesizer formats. Peptides containing N-methylated and/or α,α-disubstituted amino acid(s) are synthesized efficiently using this strategy, with crude purities of up to 98% and respectable isolated yields, greatly surpassing existing methods. It is anticipated that this approach will facilitate broader adoption of RMMR-SPPS, addressing critical technical gaps and accelerating the discovery of orally bioavailable or intracellularly targeted macrocyclic peptide drug candidates.