Oxosteroid isomerase

Enzymatic isomerization of ds-aconitate to trans-aeon itate apparently also involves proton abstraction,165 with resonance in the anion extending into the carboxylic acid group. Its mechanism may be directly related to that of the oxosteroid isomerase. However, there are other 1,3-proton shifts in which neither a carbonyl nor a carboxyl group is present in the substrate (Eqs. 13-55,13-56). 

Oxophenarsine 597s 5-Oxoproline, 551s, 662s Oxosteroid isomerase 526, 696 molecular activity 696 A5-3-Oxosteroid isomerase... 

HSD/4,5-isomerase activity [70] the action of 17/3-HSD on 4-androstenedione also provides testosterone. Inspection of Fig. 10 shows the two pathways, one involving 5-ene-3/3-hydroxysteroids (sometimes called 45), the other involving 4-en-3-oxosteroids (sometimes called the A4 pathway). It will be noted further that there are transitions from A5 to A4 at different levels via the 5-ene-3jS-HSD/isomerase enzyme system, but the reverse reactions seem to occur to only a very limited extent [52]. 

The intermediate 5-en-3-oxosteroids, such as 5-androstenedione (from DHA) are further converted by isomerization to the 4-en-3-oxosteroids by means of the 4,5-isomerase. This enzyme, which requires no cofactor, is associated with the smooth... 

Fig. 11. Proposed mechanism for the action of 3-oxosteroid 4,5-isomerase (from Talalay 1964, see Ref. 52).

Defective Bile Acid Synthesis. Specific defects in bile acid synthesis have long been postulated. Two inborn errors of bile acid synthesis, both associated with idiopathic neonatal hepatitis, A -3-oxosteroid 5-P-reductase deficiency and 3-p-hydroxy-dehydrogenase isomerase deficiency, have been described. A third disorder associated with defective bile... 

Although cholesterol is the major source of 5)9-bile acids, an unsaturated acid, 3)8-hydroxy-5-cholenic acid [174] has been found in meconium, mainly as the sulfate [175], in bile of a boy with a deficiency of 3)8-hydroxysteroid dehydrogenase [176], and in urine of healthy persons and individuals with liver disease [164]. The details of metabolism of 3)8-hydroxy-5-cholenic acid to lithocholate have not been entirely elucidated, but the mechanism for conversion of the 3/8-hydroxy-A to the 3-oxo-A derivative has been formulated in the C27 series (cf. Chapter 9). Briefly, the 3)8-ol is dehydrogenated by a microsomal enzyme fortified with NAD to provide the 3-oxo-A system [177,178]. Whether a A - A" isomerase is essential is not known, since there is no direct evidence for the formation of the intermediary 3-oxo-A system the rate-limiting step is the dehydrogenation of the 3)8-ol which may prevent accumulation of the 3-oxo-A system [177]. The reduction of the double bond at 4-5 to the 5)8- or 5a-bile acid is catalyzed by the respective A -3-oxosteroid 5)8- or 5 -reductase obtained from liver cytosol [170], and has been purified about 10-fold [178]. The formation of the 3-oxo-5/9 derivative requires the enzyme and NADPH the proton from the A side (4A-NADPH) appeared in the product as the 5)8-H, whereas the proton at C-4 is derived from the aqueous medium. Formation of the 5a derivative requires (4B-NADPH) in a similar mechanism (Fig. 4) [179], Reduction of the 3-0X0 product is then catalyzed by 3a-hydroxysteroid dehydrogenase as discussed above. 

Fig. 44.22. Biosynthesis of the adrenocorticoids from cholesterol. The enzymes involved are (a) side chain cleavage, b) 17a-hydroxylase, (c) 5-ene-3p-hydroxy-steroid dehydrogenase, (cf) 3-oxosteroid-4,5-isomerase, (e) 21-hydroxylase, (f) 11 p-hydroxylase, and (g)...

Activation of the LH receptor results in an increase of intracellular cyclic adenosine monophosphate (cAMP) levels via activation of a G protein and adenylate cyclase. In the presence of elevated concentrations of cAMP, cholesterol esterase activation occurs. This enzyme catalyzes the cleavage of cholesterol esters to free cholesterol, v/hich is then converted in mitochondria to pregnenolone as described previously. The formation of progesterone from pregnenolone is catalyzed by 5-ene-3p-hydroxysteroid dehydrogenase and 3-oxosteroid-4,5-isomerase (steps c and d in Fig. 46.3). 

These preliminary results indicate that there is a rather specific interaction between dextran-PAB and trypsin, though relatively weak compared to that observed in other experiments, in which a dextran-estradiol was used to separate 5 4 3-oxosteroid isomerase (isomerase, in the following) from a crude extract of Pseudomonas testosteroni (Table 14.2) [15]. 

In one of the earliest studies [39], we carried out the purification of A5 4 3-oxosteroid isomerase from a crude extract of Pseudomonas testosteroni by affinity pardoning in a poly(ethylene glycol)/dextran system. The macroligand was prepared by covalent coupling on poly(ethylene glycol) (PEG) of the same steroid derivative as previously described in Fig. 14.2. The resulting macroligand PEG-E,i had an inhibition constant Ki around 5 pM towards the isomerization of A5-androstene-3,17-dione, by isomerase. 

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