SEC analysis revealed that using longer chain length α,ω-ALD in the polymerization process resulted in copolymers with higher molecular weights. This was mainly attributed to the increased enzymatic reactivity and selectivity of CAL-B, which is determined by the size and shape of its active site, enabling it to accommodate substrates of specific sizes and shapes, as visualized in the proposed mechanism. It was also correlated with decreased solubility of the resulting polyesters with a shorter chain length of α,ω-ALD. Additionally, the study revealed that the melting temperatures of the copolymers were influenced by the chain length of the α,ω-ALD used in the synthesis, with longer chain length resulting in lower melting temperatures. These findings underscore the importance of considering the properties of the starting materials in the enzymatic synthesis of bio-based copolymers.