The largest proportion of commercial milk products is derived from cows worldwide. Milks from other ruminants are also important culturally and economically. Pseudomonas is frequently linked to milk spoilage under different storage temperatures. Therefore, the aim was to measure proteolytic activity of Pseudomonas spp. isolated from different raw milks under low storage temperatures. Raw milk samples of cows (n = 87), goats (n = 50), buffalos (n = 25), camels (n = 25), and yaks (n = 25) were collected from 5 provinces in China. Pseudomonas was identified by universal 16S rRNA and rpoB gene sequence analyses. Proteolytic activity on milk agar, and quantification via the trinitrobenzenesulfonic acid (TNBS) assay at 2°C, 4°C, 7°C, and 10°C were performed to ascertain proteolytic activity of Pseudomonas isolates. The TNBS can react with the released α-amino groups, indicators of protein hydrolysis, with the intensity of the yellow-orange color of reaction products. A total of 143 isolates from cow milk samples, 31 isolates from goat milks, 8 isolates from buffalo milks, 9 isolates from camel milks, and 19 isolates from yak milks were confirmed as Pseudomonas spp. Of Pseudomonas isolates, we obtained extracellular peptidase activity from 47 (22.4%) strains at 2°C, 91 (43.3%) at 4°C, 128 (61.0%) at 7°C, and 141 (67.1%) at 10°C. However, proteolytic activity of 79 (37.6%) isolates exceeded 2 μmol of glycine equivalents per mL at 10°C, followed by 61 (29.0%) at 7°C, 28 (13.3%) at 4°C, and 9 (4.3%) at 2°C. The results revealed that a large diversity of Pseudomonas spp. were present in different raw milks with the ability to produce peptidases at storage temperatures. Most isolates from cow and yak milks had high preoteolytic activity. In conclusion, our findings highlighted the importance of Pseudomonas species as broadly adaptable spoilage organisms in different types of raw milk. Low temperature and short period storage before processing (within 48 h) could reduce peptidase production of Pseudomonas spp, but milking hygiene should also be properly controlled.