Structure and Function of Human Nad+-Linked 15-Hydroxyprostaglandin Dehydrogenase
Prostaglandins are rapidly metabolized in the body. The primary route of metabolism is initiated by NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (15-PGDH) which catalyzes the reversible oxidation of 15(S)-hydroxyl group to a keto function resulting in a great loss of biological activities. 15-PGDH is therefore considered a key enzyme responsible for biological inactivation of prostaglandins. The enzyme is ubiquitously present in mammalian tissues. The first cDNA was cloned form human placenta and was found to have an open reading frame of 798 bp coding for 266 amino acids.’ The native enzyme is believed to be a dimer composed of identical subunits with a M.W. of 29 kDa. The enzyme utilizes a wide range of prostaglandins, lipoxins and hydroxy fatty acids as substrates and prefers NAD+ over NADP+ as a coenzyme.’ The enzyme is sensitive to sulfhydryl inhibitors indicating one or more cysteinyl residues are essential.’ The primary structure of 15-PGDH shows that it belongs to a superfamily of short chain dehydrogenases/reductases (SDR).3Sequence analysis indicates that an overall homology of 20% among SDRs with several conserved amino acid residues. Whether these conserved residues and cysteinyl residues are involved in catalytic activity and/or structural integrity remains to be determined. Most of these conserved residues reside in the first two thirds of the polypeptide chain suggesting that they might be involved in interacting with the common coenzyme NAD+. The C-terminal third shows little homology in sequences indicating that this segment appears to accommodate substrates of diversed structures.. This study is focused on using site-directed mutagenesis to examine the roles of several conserved and cysteinyl residues in catalytic activity and the involvement of C-terminal domain in enzyme and substrate interactions.
KeywordsSerine Prostaglandin Alanine NADH Leucine
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