Giving the imminent necessity of a new generation of biodegradable and biocompatible polymers prepared from feedstock, the synthesis of a potentially biodegradable amino acid-based copolymer by the alternating diene metathesis (ALTMET) strategy is herein presented. The reaction was tailored to minimize isomerization and deactivation of ruthenium catalysts by intramolecular coordination with the amide carbonyl group of the amino-acid-based monomer. Alternated L-lysine-phosphoester copolymers with molar masses higher than 18 000 g/mol were obtained using Hoveyda-Grubbs second-generation and Umicore M2 catalysts. The copolymer was further used to prepare nanoparticles loaded with rifampicin (up to 50 wt %) by the miniemulsion/solvent evaporation technique. The l-lysine-based copolymer is shown to be a promising material for biomedical applications, such as controlled drug delivery system.