Autophagy is a bulk degradation pathway that removes cytosolic materials to maintain cellular homeostasis. The AuTophagy-related Gene 13 (Atg13) and microtubule associate protein 1 Light Chain 3 (LC3) proteins are required for initiation of autophagy and autophagosome formation. Using crystallography and protein-protein interaction techniques we are examining the structures of human Atg13 proteins and their interaction partners, often Atg13 LIR peptides. We recently published some our work in Structure revealing that the side-chain of Lys49 of LC3 acts as a gatekeeper to regulate binding of the LIR peptide to the Atg13 proteins. Our results suggest that specific affinity of the LC3 isoforms to the Atg13 LIR peptides is required for proper autophagosome formation.