Genome-scale metabolic model (GEM) is a powerful tool for interpreting and predicting cellular phenotypes under various environmental and genetic perturbations. However, GEM only consid-ers stoichiometric constraints, and the simulated growth and product yield values will show a monotonic linear increase with increasing substrate uptake rate, which deviates from the experi-mentally measured values. Recently, the integration of enzymatic constraints into stoichiometry-based GEMs was proven to be effective in making novel discoveries and predicting new engineer-ing targets. Here we present the first genome-scale enzyme-constrained model (eciCW773) for Corynebacterium glutamicum reconstructed by integrating enzyme kinetic data from various sources using ECMpy workflow based on the high-quality GEM of C. glutamicum (obtained by modifying the iCW773 model). The enzyme-constrained model improved the prediction of pheno-types and simulated overflow metabolism, while also recapitulating the trade-off between biomass yield and enzyme usage efficiency. Finally, we used eciCW773 to identify several gene modifica-tion targets for L-lysine production, most of which agree with previously reported genes. This study shows that incorporating enzyme kinetic information into the GEM enhances the cellular phenotypes prediction of C. glutamicum, which can help identify key enzymes and thus provide reliable guidance for metabolic engineering.