AbstractMultidrug‐resistant bacteria present a major threat to public health that urgently requires new drugs or treatment approaches. Here, we conduct integrated proteomic and metabolomics analyses to screen for molecular candidates improving survival of mice infected with Vibrio parahaemolyticus, which indicate that L‐Alanine metabolism and phagocytosis are strongly correlated with mouse survival. We also assess the role of L‐Alanine in improving mouse survival by in vivo bacterial challenge experiments using various bacteria species, including V. parahaemolyticus, Escherichia coli, Pseudomonas aeruginosa, and Klebsiella pneumoniae. Functional studies demonstrate that exogenous L‐Alanine promotes phagocytosis of these multidrug‐resistant pathogen species. We reveal that the underlying mechanism involves two events boosted by L‐Alanine: TLR4 expression and L‐Alanine‐enhanced TLR4 signaling via increased biosynthesis and secretion of fatty acids, including palmitate. Palmitate enhances binding of lipopolysaccharide to TLR4, thereby promoting TLR4 dimer formation and endocytosis for subsequent activation of the PI3K/Akt and NF‐κB pathways and bacteria phagocytosis. Our data suggest that modulation of the metabolic environment is a plausible approach for combating multidrug‐resistant bacteria infection.