Aims: To investigate the diversity in specificity of cell-bound extracellular proteinases in Lactobacillus helveticus and Lactobacillus delbrueckii subsp. bulgaricus. Methods and Results: HPLC analysis of whole-cell preparations of 14 Lact. delbrueckii subsp. bulgaricus and eight Lact. helveticus strains incubated with αs1-casein (f 1–23) detected at least six distinct proteolytic patterns. Differences between groups were found in both the primary and secondary specificity toward αs1-casein (f 1–23) and its breakdown products. No correlation was found between the o-phthaldialdehyde (OPA) general proteolysis analysis and αs1-casein (f 1–23) cleavage profiles. Conclusions, Significance and Impact of Study: Using the αs1-CN (f 1–23) method, six patterns of proteolysis were found in the dairy lactobacilli tested. Understanding the influence of Lactobacillus proteinase specificity on casein degradation should facilitate efforts to develop starter cultures that predictably improve the functional properties of Mozzarella cheese.