This study aimed to obtain umami peptides from Yanjin black bone chicken and to investigate the formation mechanism of umami taste. The umami peptides were purified from the enzymatic hydrolysate of chicken using ultrafiltration (UF), gel filtration chromatography (GFC), and reversed-phase high-performance liquid chromatography (RP-HPLC). Potential novel umami peptides were then identified by nano-scale liquid chromatography-tandem mass spectrometry (nano-HPLC-MS/MS). Based on the predictions of iUmami-SCM and BIOPEP-UWM databases, five umami peptides (EELK, EEEIK, EELMK, LEEEIK, DELDKYS) with high umami scores were synthesized. Sensory evaluation revealed that these umami peptides exhibited a threshold ranging from 0.12 mg mL-1 to 0.36 mg mL-1. Circular dichroism (CD) analysis indicated the presence of β-sheet structures in the umami peptides that could be associated with taste formation. In addition, molecular docking and molecular dynamics (MD) were employed to investigate the binding mechanisms between umami peptides and the umami receptor T1R1/T1R3. The findings reveal that Lys155, Arg255, and Gln250 of T1R1/T1R3 potentially play critical roles in umami peptide binding. Taken together, our results lay a foundation for researching flavor substances and for developing flavor products from Yanjin black bone chicken.