C-Terminal region of human NAD(+)-dependent 15-hydroxyprostaglandin dehydrogenase is involved in the interaction with prostaglandin substrates

NAD(+)-dependent 15-hydroxyprostaglandin dehydrogenase (15-PGDH) catalyzes the oxidation of the 15(S) hydroxyl group of prostaglandins to a 15-keto gr oup resulting in a significant reduction of the biological activities of pr ostaglandins. Although the key residues involved in NAD(+) binding and in c atalytic activity have been partially identified, the sites of interaction of the enzyme with the prostaglandin substrates are yet to be determined. H omology analysis of the primary structures of 15-PGDH from human, mouse and rat indicates that the sequences are almost homologous except for two regi ons near the C-terminus. The involvement of the C-terminal region in cataly tic activity was examined by studies on C-terminally truncated enzymes and on human/rat chimeric enzymes. When three to four amino acids were removed successively from the C-terminal end of human 15-PGDH, the truncated enzyme s exhibited decreasing V-max/K-m ratios and increasing K-m values for PGE(2 ) as the chain was shortened. Similarly, when the C-terminal 14 amino acids of human 15-PGDH were replaced by the C-terminal 14 amino acids of rat 15- PGDH or vice versa, the V-max/K-m ratios and the K-m values for prostagland in E-2 of the chimeric enzymes were in between those of the two wild-type e nzymes. This indicates that the catalytic effectiveness of human 15-PGDH de creases as the C-terminal region is gradually removed or replaced by rat se quences. The C-terminal region appears to be more important for the interac tion of the enzyme with the prostaglandin substrates than with the coenzyme .