Activation of human placental 5-pregnene-3,20-dione isomerase activity by pyridine nucleotides.
Isomerization of 5-pregnene-3,20-dione to progesterone by human placental microsomes was stimulated by NAD and NADH. Concomitant oxidation or reduction of nucleotide was not detected based on absorbance at 340 nm. Concentrations giving half-maximum activity were 0.76 microM for NADH and 24.0 microM for NAD. Vmax values with 9.28 microM 5-pregnene-3,20-dione were 22.0 nmol/min/mg protein with NADH and 65.8 nmol/min/mg protein with NAD. When isomerase was assayed as a function of 5-pregnene-3,20-dione concentration, NAD increased Vmax but had no effect on the Km value for steroid. NADP, NADPH, acetylpyridine NAD and deamino NAD did not activate nor did they compete with NAD. Exposure of microsomes to trypsin, phospholipase A2 or phospholipase C resulted in the loss of isomerase activity. Approximately 30% of the initial activity was recovered after detergent solubilization of microsomes. Hydrogen peroxide did not affect activation by NAD. The data are consistent with nucleotide enhancement of a step in the isomerization reaction other than substrate binding.
Blomquist
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Department of Obstetrics and Gynecology, Ramsey Clinic and St Paul-Ramsey Medical Center, MN 55101.
Kotts
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Hakanson
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D003902:Detergents / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007535:Isomerases / Q000378:metabolism*; D009243:NAD / Q000494:pharmacology*; D009249:NADP / Q000494:pharmacology; D010741:Phospholipases A / Q000494:pharmacology; D054467:Phospholipases A2; D010920:Placenta / Q000201:enzymology*; D013251:Steroid Isomerases / Q000378:metabolism*; D014357:Trypsin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000494:pharmacology
D003902:Detergents; D009243:NAD; D009249:NADP; D006861:Hydrogen Peroxide; D010741:Phospholipases A; D054467:Phospholipases A2; D010738:Type C Phospholipases; D014357:Trypsin; D007535:Isomerases; D013251:Steroid Isomerases; C022660:steroid delta-isomerase