- Crystal structure of cytochrome P450 CYP105N1 from Streptomyces coelicolor, an oxidase in the coelibactin siderophore biosynthetic pathway
-
The genome sequence of Streptomyces coelicolor contains 18 cytochrome P450 enzymes. The recombinant CYP105N1 protein has been expressed in Escherichia coli and purified, and we report the biochemical and structural characterization of CYP105N1 from S. coelicolor. The purified protein exhibited the typical CO-binding spectrum of P450 enzymes and type I binding spectra with estradiol and a coelibactin analog. The oxidation of estradiol by CYP105N1, supported by H2O2, produced estriol. The crystal structure of CYP105N1 was determined at 2.9 ? resolution. An unexpected wide open binding pocket located above the heme group was identified, with a volume of approximately 4299 ?3. These results suggest that the large open pocket to the active site may be a key feature for easy access of the peptidyl carrier protein-bound substrate to perform the hydroxylation reaction. A molecular docking model with coelibactin showed that the phenyl group of coelibactin is located ? away from the heme-iron, suggesting that CYP105N1 may be involved in the hydroxylation of the phenyl ring of the coelibactin precursor during biosynthesis.
- Lim, Young-Ran,Hong, Myoung-Ki,Kim, Jin-Kwang,Doan, Thanh Thi Ngoc,Kim, Dong-Hyun,Yun, Chul-Ho,Chun, Young-Jin,Kang, Lin-Woo,Kim, Donghak
-
p. 111 - 117
(2013/02/23)
-
- STEREOSELECTIVE REDUCTION OF ALPHA-HYDROXYKETONE
-
The present invention provides methods for 1, 2-asymmetric reduction of other compounds wherein 1,2- trans diol is present. In particular the present invention discloses preparation of 17β-Estriol and its derivatives having substantially free of 17α-Estriol impurity using an achiral reductant sodium triacetoxy borohydride.
- -
-
Page/Page column 10
(2012/03/27)
-
- Estriol substantially free of 16alpha, 17alpha-estra-1,3,5(10)-triene-3,16,17-triol
-
Estriol substantially free of 16α,17α-estra-1,3,5(10)-triene-3,16,17-triol, and process for obtaining the same.
- -
-
-
- Roles of cytochromes P450 1A2 and 3A4 in the oxidation of estradiol and estrone in human liver microsomes
-
Of seven cDNA-expressed human cytochrome P450 (P450) enzymes (P450s 1A2, 2B6, 2C9, 2C19, 2D6, 2E1, and 3A4) examined, P450 1A2 was the most active in catalyzing 2- and 4-hydroxylations of estradiol and estrone. P450 3A4 and P450 2C9 also catalyzed these reactions although to lesser extents than P450 1A2. P450 1A2 also efficiently oxidized estradiol at the 16α-position but was less active in estrone 16α-hydroxylation; the latter reaction and also estradiol 16α-hydroxylation were catalyzed by P450 3A4 at significant levels. Anti-P450 1A2 antibodies inhibited 2- and 4-hydroxylations of these two estrogens catalyzed by liver microsomes of some of the human samples examined. Estradiol 16α-hydroxylation was inhibited by both anti-P450 1A2 and anti-P450 3A4, while estrone 16α-hydroxylation was significantly suppressed by anti-P450 3A4 in human liver microsomes. Fluvoxamine efficiently inhibited the estrogen hydroxylations in human liver samples that contained high levels of P450 1A2, while ketoconazole affected these activities in human samples in which P450 3A4 levels were high. α- Naphthoflavone either stimulated or had no effect on estradiol hydroxylation catalyzed by liver microsomes; the intensity of this effect depended on the human samples and their P450s. Interestingly, in the presence of anti-P450 3A4 antibodies, α-naphthoflavone was found to be able to inhibit estradiol and estrone 2-hydroxylations catalyzed by human liver microsomes. The results suggest that both P450s 1A2 and 3A4 have major roles in oxidations of estradiol and estrone in human liver and that the contents of these two P450 forms in liver microsomes determine which P450 enzymes are most important in hepatic estrogen hydroxylation by individual humans. P450 3A4 may be expected to play a more important role for some of the estrogen hydroxylation reactions than P450 1A2. Knowledge of roles of individual P450s in these estrogen hydroxylations has relevance to current controversies in hormonal carcinogenesis.
- Yamazaki, Hiroshi,Shaw, Peter M.,Guengerich, F. Peter,Shimada, Tsutomu
-
p. 659 - 665
(2007/10/03)
-
- Formylation of Oestrogens
-
Reimer-Tiemann formylations of oestradiol and oestrone were investigated and, whilst substitution was effected under certain conditions to give mixtures of 2-and 4-formyloestrogens, yields were very low and the method was unsuitable for preparative purposes.Regioselective methods were developed and 2-formyloestradiol was conveniently prepared from oestradiol by formylation of the lithio derivative of the bis(methoxymethyl) ether and removal of the protecting groups with the hydrochloric acid. 4-Formyloestradiol was prepared by a sequence of reactions starting with the methoxyethyl ether of 4-bromooestradiol, then metal-halogen interconversion, formylation with N-methylformanilide, and removal of the protecting group.A number of related derivatives, including 2-formyloestriol, were prepared.
- Pert, Derek J.,Ridley, Damon D.
-
p. 405 - 419
(2007/10/02)
-
- Mechanistic Consideration of P-450 Dependent Enzymic Reactions: Studies on Oestriol Biosynthesis
-
Methods for the synthesis of 19-hydroxy and 19-oxo derivatives of 16α-hydroxytestosterone have been developed.These compounds were labelled with 18O,2H and 3H at C-19 and also with 3H at C-17.The conversion of 3H>-16α,19-dihydroxytestosterone (11c) into oestriol (4c), using human placental aromatase was demonstrated in good yield and it was shown that in this process the 19-oxo compound (3c) is involved as an intermediate.The use of 16α,19-hydroxytestosterone, labelled with 3H predominantly in the HSi position, led to the conclusion that in oestriol biosynthesis the step, -CH2OH-> -CHO, is accompained by the loss of HRe and in the overall process the C-19 is ejected as HCOOH.On conducting experiments with either 18O2 or substrate containing 18O at C-19 it was shown that, in the conversion of 16α-hydroxy-19-oxotestosterone into oestriol, an atom of oxygen from O2 is incorporated into the formate.These features are similar to those already established for the corresponding biosynthesis of oestrone/oestradiol from androstenedione/testosterone.Our previous postulate that in oestrogen biosynthesis the same enzyme is involved in the hydroxylation reaction, -CH3 -> -CH2OH, and in the conversion, -CH2OH -> -CHO, as well as in the final cleavage of the C-10-C-19 bond is further developed.Attention is drawn to the fact that, if cytochrome P-450 dependent reactions are viewed to occur via a radical mechanism, then a concept can be developed which unifies the wide variety of transformations catalysed by this group of enzymes.The diversity of reactions would then arise from the alternative mode of decomposition of radical species by one or a combination of the following processes: (a) hydrogen abstraction; (b) disproportionation; (c) fragmentation; and (d) association of radicals.
- Stevenson, David E.,Wright, J. Neville,Akhtar, Muhammad
-
p. 2043 - 2052
(2007/10/02)
-
- Partial Purification and Characterization of Two β-Glucoronidases from Alcaligenes NG-11
-
A strain of Alcaligenes isolated from soil was a good producer of β-glucuronidase, and the enzyme was purified from the cell-free extract by sequential column chromatography on DEAE-Toyopearl, Toyopearl HW-55F, and Phenyl-Sepharose CL-4B.By these procedures, two β-glucuronidases designated as β-glucuronidases I and II were purified 240- and 508-fold, respectively. β-Glucuronidase I, with a molecular weight of 75,000, had an optimum pH at 7.5 and the enzyme II, with a molecular weight of 300,000, and maximum activity at pH 6.0.Both enzymes were strongly inhibited by saccharo-1,4-lactone, glucaro-δ-lactam, p-chloromercuribenzoate, Hg2+, and N-bromosuccinimide. β-Glucuronidase I was active toward estrogen-3-β-glucuronides and inert toward β-glucuronide conjugates of menthol, estrogen-17β-, estrogen-16α-, androsterone-3α-, testosterone-17β-, cortisol-17α-. β-Glucuronidase II hydrolyzed all of these substrates. β-Glucuronidase I was inhhibited by phenolphthalein and its glucuronide.
- Ogushi, Susumu,Koga, Satoshi,Ito, Kiyoshi,Makino, Yasutaka,Ando, Makoto,Tsuru, Daisuke
-
p. 3093 - 3100
(2007/10/02)
-
- Synthesis of 2-Methoxy- and 4-Methoxy-Estrogens with Halogen-Methoxy Exchange Reaction
-
Synthesis of 2-methoxy- and 4-methoxy-estrone (6) and (9), 2-methoxy- and 4-methoxy-estradiol (15) and (18), and 2-methoxy- and 4-methoxy-estratriol (24) and (27) are described.Catalytic hydrogenation over Pd/C of 2,4-dibromo or 2,4-diiodo estrogens gave regioselectively the corresponding 4-halogeno derivatives in excellent yields.Reaction of 2-iodo or 4-iodo estradiol and 2-iodo or 4-iodo estriol with NaOCH3 in MeOH and dimethylformamide (DMF) in the presence of CuCl2 gave in an excellent yield and in a good yield, while (6) and (9) were also similarly obtained by the reaction with pyridine instead of DMF.
- Numazawa, Mitsuteri,Ogura, Yuko,Kimura, Katsuhiko,Nagaoka, Masao
-
p. 3701 - 3715
(2007/10/02)
-
- Novel and Efficient Synthesis of Estriol and its 16-Glucuronide via 2,4,16α-Tribromoestrone
-
A novel synthesis of estra-1,3,5(10)-triene-3,16α,17β-triol (5), sodium 3,17β-dihydroxyestra-1,3,5(10)-trien-16α-yl-β-D-glucopyranosuronate (12b) aand sodium 3-hydroxyestra-1,3,5(10)-trien-17β-yl-β-D-glucopyranosuronate (10b) is described. 2,4,16α-Tribromo-3-hydroxyestra-1,3,5(10)-trien-17-one (2a) was efficiently synthesized in one step with quantitative yield by bromination of 3-hydroxyestra-1,3,5(10)-trien-17-one (1) with cupric bromide.Treatment of (2a) with NaOH in aqueous pyridine under the controlled conditions gave the 16α-hydroxy-17-ketone (4a) without ketol rearrangement.The ketol (4a) was converted in quantitative yield into the triol (5) via a sodium borohydride reduction in the presence of palladium chloride.Reaction of (4a) with methyl 1-bromo-1-deoxy-2,3,4-tri-O-acetyl-α-D-glucopyranosuronate using silver carbonate as a catalyst yielded the 16-monoglucuronide acetate methyl ester (11).The reductive removal of the bromines of (11) with sodium borohydride followed by NaOH hydrolysis gave the glucuronide (12b).A direct glucuronidation of 2,4-dibromoestra-1,3,5(10)-triene-3,17β-diol (8) and a subsequent hydrolysis of the 17-glucuronide (9) gave the glucuronide (10b).
- Numazawa, Mitsuteru,Nagaoka, Masao,Tsuji, Masachika,Osawa, Yoshio
-
p. 121 - 125
(2007/10/02)
-
- IDENTIFICATION OF 6α- AND 7α-HYDROXYESTRONE AS MAJOR METABOLITES OF ESTRONE AND ESTRADIOL IN PORCINE UTERUS
-
Polar metabolites extracted from the effluents of viable porcine uterine strips superfused with either 6,7-3H-estrone or 6,7-3H-estradiol were identified as a 1 : 1 mixture of 6α-hydroxyestrone and 7α-hydroxyestrone by paper chromatography in various systems, derivatization and crystallizations to a constant specific activity.The hydroxylated compounds are the only derivatives detected after estrone superfusion.The major metabolite of estradiol released in short-time experiments is estrone followed by its 6α- and 7α-hydroxylated derivatives.
- Maschler, I.,Ball, P.,Bayerkoehler, G.,Gaues, J.,Knuppen, R.
-
p. 597 - 608
(2007/10/02)
-
- Novel N-nitroso compounds, compositions containing such compounds, processes for their preparation and methods of treatment therewith, and novel intermediates
-
This invention relates to novel N-halogenoalkyl-N-nitroso carbamates and N 4 halogenalkyl-N 4 -nitroso allophanates of steroid compounds, having an anti-tumor activity, and to the preparation thereof. The invention is also concerned with pharmaceutical compositions containing the said compounds, and methods of treatment therewith.
- -
-
-
