Arsenic is a class I human carcinogen (such as inducing skin cancer) by its prominent chemical interaction with protein thio (-SH) group. Therefore, arsenic may compromise protein S-nitrosylation by competing the -SH binding activity. In the present study, we aimed to understand the influence of arsenic on protein S-nitrosylation and the following proteomic changes. By using primary human skin keratinocyte, we found that arsenic treatment decreased the level of protein S-nitrosylation. This was coincident to the decent expressions of endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS). By using LC-MS/MS, around twenty S-nitrosoproteins were detected in the biotin-switched eluent. With the interest that arsenic not only regulates posttranslational S-nitrosylation but also separately affects protein's translation expression, we performed two-dimensional gel electrophoresis and found that 8 proteins were significantly decreased during arsenic treatment. Whether these decreased proteins are the consequence of protein S-nitrosylation will be further investigated. Taken together, these results provide a finding that arsenic can deplete the binding activity of NO and therefore reduce protein S-nitrosylation.