The polymer was only partially soluble in the organic solvent. Several other solvents (DMF, DMSO, CH2Cl2, CHCl3) were employed, but none achieved complete dissolution of the copolymer precipitate. It is hypothesised that cross-linking happened during the chain-extension, possibly due to transesterification of the side chains once inaccessible to the catalyst, which rendered part of the final copolymer insoluble. The fraction of insoluble copolymer was 62% ± 8% irrespective of the molecular weight of the formed polymers or the morphology of the self-assemblies. The soluble parts of the bioPISA-derived diblock copolymers had number average molecular weights (Mn) of 30-60 kDa and dispersities (Ð) of 1.5-2.5 (Figure and Table ). One explanation for the broad molecular weight distributions is that, unlike bioRAFT where the polymer chain does not need proximity to the enzyme to grow, ATRP necessitates the catalyst for the atom transfer (in this case Mb) to be close to the active polymer chain-end. In PISA, the growing chain-ends are located in the hydrophobic core or membrane of the self-assemblies, and therefore difficult for the water-soluble enzyme to reach. This most likely led to a reduction in the degree of control of the polymerization and, as a result, in high dispersities. Thus, most likely the reaction proceeded through an ATRP-like initiation and initial RDRP until the block copolymers become water-insoluble, followed by a free radical polymerisation inside of the hydrophobic parts of the formed self-assemblies. While this might seem to be a disadvantage, it did not significantly affect the predictability of our method in generating a variety of self-assembled nanostructures, as discussed above.