Chlorate is known to be an in vitro inhibitor of ATP-sulfurylase, the first enzyme in the biosynthesis of PAPS which is the ubiquitous co-substrate for sulfation. Here, the effect of chlorate on protein sulfation in intact cells was investigated. Treatment of various cell cultures with 1 mM sodium chlorate in a medium low in sulfate and sulfur-containing amino acids resulted in an inhibition of protein sulfation greater than 95%. Tyrosine as well as carbohydrate sulfation was blocked. Chlorate did not inhibit protein synthesis and did not exhibit any other toxic effects, even after prolonged treatment of cell cultures. Thus, chlorate treatment provides a powerful tool for studying the biological significance of protein sulfation.