To further explore the tolerance and scope of this cyclisation methodology we synthesised several pentapeptides carrying a range of proteinogenic amino acids (14-20; Figure ). When positioning the alGly residue at the N-terminus and the Cys at the C-terminus, effective N-to C-terminal macrocyclisation was realised for all sequences explored to yield 17mer macrocycles (wrt the number of bonds within the macrocycle) in moderate to excellent yields (21-27, 48-74% isolated yield). The model peptides explored carried the majority of the 20 canonical amino acids, demonstrating that this reaction can tolerate the presence of the diverse chemical functionality displayed across the proteome. Interestingly, switching the terminal residues eliminated or hindered conversion to the desired product for this size of macrocycle (Table ). To explore macrocyclisation of a peptide carrying the radical trap residue at an internal position, peptide 20 was synthesised with alGly as the penultimate N-terminal residue. For this model, cyclisation was successful in high yield (27, 82%). Moving to longer sequences, we explored several 6, 7, & 8mer peptides carrying a range of proteinogenic residues (28-38) to yield 20, 23, and 26mer peptide macrocycles in moderate to excellent yield (39-49, 32-76%, Figure ). All macrocyclic products were characterised by analytical HPLC, MS and 1 H NMR; CD spectroscopy was run for examples of each size of macrocycle (SI). Switching the N-and C-terminal residues for model 29 (H-CKISY(alG)-NH2) had no effect on yield for this size of macrocycle, unlike the smaller model peptides explored (Table ). In several cases of low or negligible yield (e.g., 41a) acetylation of the N-terminus reinstated a moderate yield (41b, 48%). To confirm the effect that buffer composition has on the isolated yield, cyclisation of model 28 was carried out in PBS. Conversion to the desired product was again not as effective in this buffer compared to cyclisation in conjugation buffer. However, for this model, the ratio of product to desulfurised linear by-product was comparable to that of the conjugation buffer example. Additionally, PTA was revisited for the low-yielding model 32. While this additive did not give an improvement in yield for model 7a, we found that it improved the conversion to the product for the more complex model 32, increasing the yield from 37% to 52%). Therefore, while TCEP gives more favourable results overall, alternative phosphines can be employed.