Data in this article are associated with the research article “High-throughput screening of enzyme mutants by comparison of their activity ratios to an enzyme tag” (Li et al., 2019) [1]. Data are provided on the development of a system for high-throughput (HTP) screening of mutants through the comparison of the activity ratios of an applicable enzyme and its mutants to a suitable tag enzyme in cell lysates of their fused forms, with Escherichia coli alkaline phosphatase (ECAP) as the tag fused to the N-terminus of Pseudomonas Aeruginosa arylsulfatase (PAAS) and its mutants via a flexible linker. Data were made publicly available for further analyses.