Schizosaccharomyces pombe Fcp1 is an essential protein serine phosphatase that preferentially dephosphorylates Ser(2) of the RNA polymerase II C-terminal domain (CTD) heptad repeat Y(1)S(2)P(3)T(4)S(5)P(6)S(7). Here we show that: (i) Fcp1 acts distributively during the hydrolysis of substrates containing tandem Ser(2)-PO(4) heptads; (ii) the minimal optimal CTD substrate for Fcp1 is a single heptad of phasing S(5)P(6)S(7)Y(1)S(2)P(3)T(4); and (iii) single alanine mutations of flanking residues Tyr(1) or Pro(3) result in 6-fold decrements in CTD phosphatase activity. Fcp1 belongs to the DXDX(T/V) family of phosphotransferases that act via an acyl-phosphoenzyme intermediate. An alanine scan of 11 conserved positions of S. pombe Fcp1 identifies Thr(174), Tyr(237), Thr(243), and Tyr(249) as important for phosphatase activity. Structure-activity relationships at these positions were determined by introducing conservative substitutions. Our results, together with previous mutational studies, highlight a constellation of 11 amino acids that are conserved in all Fcp1 orthologs and likely comprise the active site.