Metal corrosion can cause massive economic losses and many safety hazards; thus, metal corrosion inhibition has been a major research direction worldwide. Among the many methods to stop metal corrosion, the addition of corrosion inhibitors is a common method. The use of amino acids as metal corrosion inhibitors not only has the advantage of being economical and efficient but also meets the long-term concept of being environmentally friendly. In this study, the effect on the corrosion behavior of a mild steel (X100 pipeline steel) in 1 M HCl containing the same concentration of the amino groups of cystine (Cys-Cys), cysteine (Cys) and glutamate (Glu) was investigated via density functional theory (DFT) and various characterization methods. These methods include the weightlessness method, electrochemical tests, scanning electron microscopy (SEM), and the contact angle test. The results show that the inhibition efficiency increased with increasing inhibitor concentration; Cys-Cys showed the best inhibition and Glu showed the poorest inhibition for X100 pipeline steel at the same amino group concentration. Furthermore, the results obtained from various characterization methods were generally consistent.