Aims To identify the differences that account for the acid sensitivity of Lactobacillus casei Delta RR12. RR12 controls the expression of the dlt operon, and its inactivation leads to a diminished teichoic acid D-alanylation activity. To this end, a comparison of its response of Delta RR12 to low pH with the parental strain Lact.casei BL23 was carried out. Methods and Results The ability to induce an acid tolerance response (ATR), fatty acid (FA) composition and proteome changes induced in both strains in response to acid were investigated. Results obtained showed that both strains induce a growth-phase-dependent ATR. However, significant differences in the content of FAs and membrane-associated proteins were detected. Conclusions The greater abundance of cytoplasmic proteins in the membrane fraction of the mutant strain Delta RR12 suggests an increased permeability of the cell membrane in this strain. Significance and Impact of the Study The analysis of the response to low pH of strain Delta RR12 indicated that the inactivation of TCS12 affected the content of FAs and proteins associated to the cell envelope. Increased abundance of cytoplasmic proteins suggested that low alanylation of teichoic acids affected the permeability of the cell membrane and possibly accounts for the acid sensitivity of strain Delta RR12.