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Stefan A Wudy - One of the best experts on this subject based on the ideXlab platform.
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role of steroid sulfatase in steroid homeostasis and characterization of the Sulfated steroid pathway evidence from steroid sulfatase deficiency
Molecular and Cellular Endocrinology, 2016Co-Authors: Alberto Sanchezguijo, Rita Bernhardt, Jens Neunzig, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Adrian Gerber, Hanschristian Schuppe, Stefan A WudyAbstract:The impact of steroid sulfatase (STS) activity in the circulating levels of both Sulfated and unconjugated steroids is only partially known. In addition, the Sulfated steroid pathway, a parallel pathway to the one for unconjugated steroids, which uses the same enzymes, has never been characterized in detail before. Patients with steroid sulfatase deficiency (STSD) are unable to enzymatically convert Sulfated steroids into their unconjugated forms, and are a good model to elucidate how STS affects steroid biosynthesis and to study the metabolism of Sulfated steroids. We quantified unconjugated and Sulfated steroids in STSD serum, and compared these results with data obtained from serum of healthy controls. Most Sulfated steroids were increased in STSD. However, androstenediol-3-Sulfate and epiandrosterone Sulfate showed similar levels in both groups, and the concentrations of androsterone Sulfate were notably lower. Hydroxylated forms of DHEAS and of pregnenolone Sulfate were found to be increased in STSD, suggesting a mechanism to improve the excretion of Sulfated steroids. STSD testosterone concentrations were normal, but cholesterol and DHEA were significantly decreased. Additionally, serum bile acids were three-fold higher in STSD. Correlations between concentrations of steroids in each group indicate that 17α-hydroxy-pregnenolone-3-Sulfate in men is mainly biosynthesized from the precursor pregnenolone Sulfate and androstenediol-3-Sulfate from DHEAS. These findings confirm the coexistence of two steroidogenic pathways: one for unconjugated steroids and another one for Sulfated steroids. Each pathway is responsible for the synthesis of specific steroids. The equal levels of testosterone, and the reduced level of unconjugated precursors in STSD, support that testosterone is primarily synthesized from Sulfated steroids. In consequence, testosterone synthesis in STSD relies on an enzyme with sulfatase activity other than STS. This study reveals that STS is a key player of steroid biosynthesis regulating the availability of circulating cholesterol.
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Stefan A WudyAbstract:Steroids are primarily present in human fl uids in their Sulfated forms. Profi ling of these compounds is impor- tant from both diagnostic and physiological points of view . Here, we present a novel method for the quantifi cation of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quan- tifi ed for the fi rst time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16- -hydroxy-dehydroepiandrosterone Sulfate, dehydroepi- androsterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epites- tosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 l of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound accord- ing to its physiological concentration. The assay showed good linearity (R 2 > 0.99) and recovery for all the compounds, with limits of quantifi cation ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coeffi cient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the sul- fated steroidome in diseases such as steroid sulfatase defi - ciency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantifi cation of Sulfated steroids in human blood. — S a�nchez-Guijo, A., V. Oji, M. F. Hartmann, H. Traupe, and S. A. Wudy. Simultaneous quantifi cation of cholesterol sul- fate, androgen Sulfates, and progestagen Sulfates in human serum by LC-MS/MS. J. Lipid Res. 2015. 56: 1843-1851. Abbreviations: ACN, acetonitrile; AnDiolS, 5-androsten-3 ,17 - diol-3-Sulfate (androstenediol Sulfate); AnS, 5 -androstan-3 -ol-17- one-3-Sulfate (androsterone Sulfate); CS, 5-cholesten-3 -ol-3-Sulfate (cholesterol Sulfate); CV, coeffi cient of variation; DHEAS, 5-androsten- 3 -ol-17-one-3-Sulfate (dehydroepiandrosterone Sulfate); DHTS, 5 - androstan-17 -ol-3-one-17-Sulfate (dihydrotestosterone Sulfate); epiAnS, 5 -androstan-3 -ol-17-one-3-Sulfate (epiandrosterone Sulfate); E1S, estrone Sulfate; E2S, estradiol Sulfate; E3S, estriol Sulfate; eTS, 4- androsten-17 -ol-3-one-17-Sulfate (epitestosterone Sulfate); IS, inter- nal standard; LOD, limit of detection; LOQ, limit of quantifi cation; MeOH, methanol; 16OHDHEAS, 5-androsten-3 ,16 -diol-17-one-3- Sulfate (16- -hydroxy-dehydroepiandrosterone Sulfate); 17OHPregS, 5-pregnen-3 ,17 -diol-20-one-3-Sulfate (17-hydroxy-pregnenolone Sulfate); PregS, 5-pregnen-3 -ol-20-one-3-Sulfate; QC, quality control sample; RE, relative error; RXLI, recessive X-linked ichthyosis; STS, steroid sulfatase; TS, 4-androsten-17 -ol-3-one-17-Sulfate (testosterone Sulfate ).
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Stefan A WudyAbstract:Steroids are primarily present in human fluids in their Sulfated forms. Profiling of these compounds is important from both diagnostic and physiological points of view. Here, we present a novel method for the quantification of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quantified for the first time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16-α-hydroxy-dehydroepiandrosterone Sulfate, dehydroepiandrosterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epitestosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 μl of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound according to its physiological concentration. The assay showed good linearity (R(2) > 0.99) and recovery for all the compounds, with limits of quantification ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coefficient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the Sulfated steroidome in diseases such as steroid sulfatase deficiency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantification of Sulfated steroids in human blood.
Alberto Sanchezguijo - One of the best experts on this subject based on the ideXlab platform.
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role of steroid sulfatase in steroid homeostasis and characterization of the Sulfated steroid pathway evidence from steroid sulfatase deficiency
Molecular and Cellular Endocrinology, 2016Co-Authors: Alberto Sanchezguijo, Rita Bernhardt, Jens Neunzig, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Adrian Gerber, Hanschristian Schuppe, Stefan A WudyAbstract:The impact of steroid sulfatase (STS) activity in the circulating levels of both Sulfated and unconjugated steroids is only partially known. In addition, the Sulfated steroid pathway, a parallel pathway to the one for unconjugated steroids, which uses the same enzymes, has never been characterized in detail before. Patients with steroid sulfatase deficiency (STSD) are unable to enzymatically convert Sulfated steroids into their unconjugated forms, and are a good model to elucidate how STS affects steroid biosynthesis and to study the metabolism of Sulfated steroids. We quantified unconjugated and Sulfated steroids in STSD serum, and compared these results with data obtained from serum of healthy controls. Most Sulfated steroids were increased in STSD. However, androstenediol-3-Sulfate and epiandrosterone Sulfate showed similar levels in both groups, and the concentrations of androsterone Sulfate were notably lower. Hydroxylated forms of DHEAS and of pregnenolone Sulfate were found to be increased in STSD, suggesting a mechanism to improve the excretion of Sulfated steroids. STSD testosterone concentrations were normal, but cholesterol and DHEA were significantly decreased. Additionally, serum bile acids were three-fold higher in STSD. Correlations between concentrations of steroids in each group indicate that 17α-hydroxy-pregnenolone-3-Sulfate in men is mainly biosynthesized from the precursor pregnenolone Sulfate and androstenediol-3-Sulfate from DHEAS. These findings confirm the coexistence of two steroidogenic pathways: one for unconjugated steroids and another one for Sulfated steroids. Each pathway is responsible for the synthesis of specific steroids. The equal levels of testosterone, and the reduced level of unconjugated precursors in STSD, support that testosterone is primarily synthesized from Sulfated steroids. In consequence, testosterone synthesis in STSD relies on an enzyme with sulfatase activity other than STS. This study reveals that STS is a key player of steroid biosynthesis regulating the availability of circulating cholesterol.
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Stefan A WudyAbstract:Steroids are primarily present in human fl uids in their Sulfated forms. Profi ling of these compounds is impor- tant from both diagnostic and physiological points of view . Here, we present a novel method for the quantifi cation of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quan- tifi ed for the fi rst time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16- -hydroxy-dehydroepiandrosterone Sulfate, dehydroepi- androsterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epites- tosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 l of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound accord- ing to its physiological concentration. The assay showed good linearity (R 2 > 0.99) and recovery for all the compounds, with limits of quantifi cation ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coeffi cient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the sul- fated steroidome in diseases such as steroid sulfatase defi - ciency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantifi cation of Sulfated steroids in human blood. — S a�nchez-Guijo, A., V. Oji, M. F. Hartmann, H. Traupe, and S. A. Wudy. Simultaneous quantifi cation of cholesterol sul- fate, androgen Sulfates, and progestagen Sulfates in human serum by LC-MS/MS. J. Lipid Res. 2015. 56: 1843-1851. Abbreviations: ACN, acetonitrile; AnDiolS, 5-androsten-3 ,17 - diol-3-Sulfate (androstenediol Sulfate); AnS, 5 -androstan-3 -ol-17- one-3-Sulfate (androsterone Sulfate); CS, 5-cholesten-3 -ol-3-Sulfate (cholesterol Sulfate); CV, coeffi cient of variation; DHEAS, 5-androsten- 3 -ol-17-one-3-Sulfate (dehydroepiandrosterone Sulfate); DHTS, 5 - androstan-17 -ol-3-one-17-Sulfate (dihydrotestosterone Sulfate); epiAnS, 5 -androstan-3 -ol-17-one-3-Sulfate (epiandrosterone Sulfate); E1S, estrone Sulfate; E2S, estradiol Sulfate; E3S, estriol Sulfate; eTS, 4- androsten-17 -ol-3-one-17-Sulfate (epitestosterone Sulfate); IS, inter- nal standard; LOD, limit of detection; LOQ, limit of quantifi cation; MeOH, methanol; 16OHDHEAS, 5-androsten-3 ,16 -diol-17-one-3- Sulfate (16- -hydroxy-dehydroepiandrosterone Sulfate); 17OHPregS, 5-pregnen-3 ,17 -diol-20-one-3-Sulfate (17-hydroxy-pregnenolone Sulfate); PregS, 5-pregnen-3 -ol-20-one-3-Sulfate; QC, quality control sample; RE, relative error; RXLI, recessive X-linked ichthyosis; STS, steroid sulfatase; TS, 4-androsten-17 -ol-3-one-17-Sulfate (testosterone Sulfate ).
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Stefan A WudyAbstract:Steroids are primarily present in human fluids in their Sulfated forms. Profiling of these compounds is important from both diagnostic and physiological points of view. Here, we present a novel method for the quantification of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quantified for the first time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16-α-hydroxy-dehydroepiandrosterone Sulfate, dehydroepiandrosterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epitestosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 μl of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound according to its physiological concentration. The assay showed good linearity (R(2) > 0.99) and recovery for all the compounds, with limits of quantification ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coefficient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the Sulfated steroidome in diseases such as steroid sulfatase deficiency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantification of Sulfated steroids in human blood.
Michaela F Hartmann - One of the best experts on this subject based on the ideXlab platform.
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role of steroid sulfatase in steroid homeostasis and characterization of the Sulfated steroid pathway evidence from steroid sulfatase deficiency
Molecular and Cellular Endocrinology, 2016Co-Authors: Alberto Sanchezguijo, Rita Bernhardt, Jens Neunzig, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Adrian Gerber, Hanschristian Schuppe, Stefan A WudyAbstract:The impact of steroid sulfatase (STS) activity in the circulating levels of both Sulfated and unconjugated steroids is only partially known. In addition, the Sulfated steroid pathway, a parallel pathway to the one for unconjugated steroids, which uses the same enzymes, has never been characterized in detail before. Patients with steroid sulfatase deficiency (STSD) are unable to enzymatically convert Sulfated steroids into their unconjugated forms, and are a good model to elucidate how STS affects steroid biosynthesis and to study the metabolism of Sulfated steroids. We quantified unconjugated and Sulfated steroids in STSD serum, and compared these results with data obtained from serum of healthy controls. Most Sulfated steroids were increased in STSD. However, androstenediol-3-Sulfate and epiandrosterone Sulfate showed similar levels in both groups, and the concentrations of androsterone Sulfate were notably lower. Hydroxylated forms of DHEAS and of pregnenolone Sulfate were found to be increased in STSD, suggesting a mechanism to improve the excretion of Sulfated steroids. STSD testosterone concentrations were normal, but cholesterol and DHEA were significantly decreased. Additionally, serum bile acids were three-fold higher in STSD. Correlations between concentrations of steroids in each group indicate that 17α-hydroxy-pregnenolone-3-Sulfate in men is mainly biosynthesized from the precursor pregnenolone Sulfate and androstenediol-3-Sulfate from DHEAS. These findings confirm the coexistence of two steroidogenic pathways: one for unconjugated steroids and another one for Sulfated steroids. Each pathway is responsible for the synthesis of specific steroids. The equal levels of testosterone, and the reduced level of unconjugated precursors in STSD, support that testosterone is primarily synthesized from Sulfated steroids. In consequence, testosterone synthesis in STSD relies on an enzyme with sulfatase activity other than STS. This study reveals that STS is a key player of steroid biosynthesis regulating the availability of circulating cholesterol.
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Stefan A WudyAbstract:Steroids are primarily present in human fl uids in their Sulfated forms. Profi ling of these compounds is impor- tant from both diagnostic and physiological points of view . Here, we present a novel method for the quantifi cation of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quan- tifi ed for the fi rst time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16- -hydroxy-dehydroepiandrosterone Sulfate, dehydroepi- androsterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epites- tosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 l of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound accord- ing to its physiological concentration. The assay showed good linearity (R 2 > 0.99) and recovery for all the compounds, with limits of quantifi cation ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coeffi cient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the sul- fated steroidome in diseases such as steroid sulfatase defi - ciency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantifi cation of Sulfated steroids in human blood. — S a�nchez-Guijo, A., V. Oji, M. F. Hartmann, H. Traupe, and S. A. Wudy. Simultaneous quantifi cation of cholesterol sul- fate, androgen Sulfates, and progestagen Sulfates in human serum by LC-MS/MS. J. Lipid Res. 2015. 56: 1843-1851. Abbreviations: ACN, acetonitrile; AnDiolS, 5-androsten-3 ,17 - diol-3-Sulfate (androstenediol Sulfate); AnS, 5 -androstan-3 -ol-17- one-3-Sulfate (androsterone Sulfate); CS, 5-cholesten-3 -ol-3-Sulfate (cholesterol Sulfate); CV, coeffi cient of variation; DHEAS, 5-androsten- 3 -ol-17-one-3-Sulfate (dehydroepiandrosterone Sulfate); DHTS, 5 - androstan-17 -ol-3-one-17-Sulfate (dihydrotestosterone Sulfate); epiAnS, 5 -androstan-3 -ol-17-one-3-Sulfate (epiandrosterone Sulfate); E1S, estrone Sulfate; E2S, estradiol Sulfate; E3S, estriol Sulfate; eTS, 4- androsten-17 -ol-3-one-17-Sulfate (epitestosterone Sulfate); IS, inter- nal standard; LOD, limit of detection; LOQ, limit of quantifi cation; MeOH, methanol; 16OHDHEAS, 5-androsten-3 ,16 -diol-17-one-3- Sulfate (16- -hydroxy-dehydroepiandrosterone Sulfate); 17OHPregS, 5-pregnen-3 ,17 -diol-20-one-3-Sulfate (17-hydroxy-pregnenolone Sulfate); PregS, 5-pregnen-3 -ol-20-one-3-Sulfate; QC, quality control sample; RE, relative error; RXLI, recessive X-linked ichthyosis; STS, steroid sulfatase; TS, 4-androsten-17 -ol-3-one-17-Sulfate (testosterone Sulfate ).
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Stefan A WudyAbstract:Steroids are primarily present in human fluids in their Sulfated forms. Profiling of these compounds is important from both diagnostic and physiological points of view. Here, we present a novel method for the quantification of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quantified for the first time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16-α-hydroxy-dehydroepiandrosterone Sulfate, dehydroepiandrosterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epitestosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 μl of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound according to its physiological concentration. The assay showed good linearity (R(2) > 0.99) and recovery for all the compounds, with limits of quantification ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coefficient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the Sulfated steroidome in diseases such as steroid sulfatase deficiency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantification of Sulfated steroids in human blood.
Heiko Traupe - One of the best experts on this subject based on the ideXlab platform.
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role of steroid sulfatase in steroid homeostasis and characterization of the Sulfated steroid pathway evidence from steroid sulfatase deficiency
Molecular and Cellular Endocrinology, 2016Co-Authors: Alberto Sanchezguijo, Rita Bernhardt, Jens Neunzig, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Adrian Gerber, Hanschristian Schuppe, Stefan A WudyAbstract:The impact of steroid sulfatase (STS) activity in the circulating levels of both Sulfated and unconjugated steroids is only partially known. In addition, the Sulfated steroid pathway, a parallel pathway to the one for unconjugated steroids, which uses the same enzymes, has never been characterized in detail before. Patients with steroid sulfatase deficiency (STSD) are unable to enzymatically convert Sulfated steroids into their unconjugated forms, and are a good model to elucidate how STS affects steroid biosynthesis and to study the metabolism of Sulfated steroids. We quantified unconjugated and Sulfated steroids in STSD serum, and compared these results with data obtained from serum of healthy controls. Most Sulfated steroids were increased in STSD. However, androstenediol-3-Sulfate and epiandrosterone Sulfate showed similar levels in both groups, and the concentrations of androsterone Sulfate were notably lower. Hydroxylated forms of DHEAS and of pregnenolone Sulfate were found to be increased in STSD, suggesting a mechanism to improve the excretion of Sulfated steroids. STSD testosterone concentrations were normal, but cholesterol and DHEA were significantly decreased. Additionally, serum bile acids were three-fold higher in STSD. Correlations between concentrations of steroids in each group indicate that 17α-hydroxy-pregnenolone-3-Sulfate in men is mainly biosynthesized from the precursor pregnenolone Sulfate and androstenediol-3-Sulfate from DHEAS. These findings confirm the coexistence of two steroidogenic pathways: one for unconjugated steroids and another one for Sulfated steroids. Each pathway is responsible for the synthesis of specific steroids. The equal levels of testosterone, and the reduced level of unconjugated precursors in STSD, support that testosterone is primarily synthesized from Sulfated steroids. In consequence, testosterone synthesis in STSD relies on an enzyme with sulfatase activity other than STS. This study reveals that STS is a key player of steroid biosynthesis regulating the availability of circulating cholesterol.
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Stefan A WudyAbstract:Steroids are primarily present in human fl uids in their Sulfated forms. Profi ling of these compounds is impor- tant from both diagnostic and physiological points of view . Here, we present a novel method for the quantifi cation of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quan- tifi ed for the fi rst time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16- -hydroxy-dehydroepiandrosterone Sulfate, dehydroepi- androsterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epites- tosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 l of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound accord- ing to its physiological concentration. The assay showed good linearity (R 2 > 0.99) and recovery for all the compounds, with limits of quantifi cation ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coeffi cient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the sul- fated steroidome in diseases such as steroid sulfatase defi - ciency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantifi cation of Sulfated steroids in human blood. — S a�nchez-Guijo, A., V. Oji, M. F. Hartmann, H. Traupe, and S. A. Wudy. Simultaneous quantifi cation of cholesterol sul- fate, androgen Sulfates, and progestagen Sulfates in human serum by LC-MS/MS. J. Lipid Res. 2015. 56: 1843-1851. Abbreviations: ACN, acetonitrile; AnDiolS, 5-androsten-3 ,17 - diol-3-Sulfate (androstenediol Sulfate); AnS, 5 -androstan-3 -ol-17- one-3-Sulfate (androsterone Sulfate); CS, 5-cholesten-3 -ol-3-Sulfate (cholesterol Sulfate); CV, coeffi cient of variation; DHEAS, 5-androsten- 3 -ol-17-one-3-Sulfate (dehydroepiandrosterone Sulfate); DHTS, 5 - androstan-17 -ol-3-one-17-Sulfate (dihydrotestosterone Sulfate); epiAnS, 5 -androstan-3 -ol-17-one-3-Sulfate (epiandrosterone Sulfate); E1S, estrone Sulfate; E2S, estradiol Sulfate; E3S, estriol Sulfate; eTS, 4- androsten-17 -ol-3-one-17-Sulfate (epitestosterone Sulfate); IS, inter- nal standard; LOD, limit of detection; LOQ, limit of quantifi cation; MeOH, methanol; 16OHDHEAS, 5-androsten-3 ,16 -diol-17-one-3- Sulfate (16- -hydroxy-dehydroepiandrosterone Sulfate); 17OHPregS, 5-pregnen-3 ,17 -diol-20-one-3-Sulfate (17-hydroxy-pregnenolone Sulfate); PregS, 5-pregnen-3 -ol-20-one-3-Sulfate; QC, quality control sample; RE, relative error; RXLI, recessive X-linked ichthyosis; STS, steroid sulfatase; TS, 4-androsten-17 -ol-3-one-17-Sulfate (testosterone Sulfate ).
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simultaneous quantification of cholesterol Sulfate androgen Sulfates and progestagen Sulfates in human serum by lc ms ms
Journal of Lipid Research, 2015Co-Authors: Alberto Sanchezguijo, Michaela F Hartmann, Heiko Traupe, Vinzenz Oji, Stefan A WudyAbstract:Steroids are primarily present in human fluids in their Sulfated forms. Profiling of these compounds is important from both diagnostic and physiological points of view. Here, we present a novel method for the quantification of 11 intact steroid Sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quantified for the first time in blood, include cholesterol Sulfate, pregnenolone Sulfate, 17-hydroxy-pregnenolone Sulfate, 16-α-hydroxy-dehydroepiandrosterone Sulfate, dehydroepiandrosterone Sulfate, androstenediol Sulfate, androsterone Sulfate, epiandrosterone Sulfate, testosterone Sulfate, epitestosterone Sulfate, and dihydrotestosterone Sulfate. The assay was conceived to quantify Sulfated steroids in a broad range of concentrations, requiring only 300 μl of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound according to its physiological concentration. The assay showed good linearity (R(2) > 0.99) and recovery for all the compounds, with limits of quantification ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coefficient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the Sulfated steroidome in diseases such as steroid sulfatase deficiency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantification of Sulfated steroids in human blood.
Bert Van Loo - One of the best experts on this subject based on the ideXlab platform.
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structural and mechanistic analysis of the choline sulfatase from sinorhizobium melliloti a class i sulfatase specific for an alkyl Sulfate ester
Journal of Molecular Biology, 2018Co-Authors: Bert Van Loo, Markus Schober, Eugene Valkov, Magdalena Heberlein, Erich Bornbergbauer, Kurt Faber, M Hyvonen, Florian HollfelderAbstract:Abstract Hydrolysis of organic Sulfate esters proceeds by two distinct mechanisms, water attacking at either sulfur (S–O bond cleavage) or carbon (C–O bond cleavage). In primary and secondary alkyl Sulfates, attack at carbon is favored, whereas in aromatic Sulfates and Sulfated sugars, attack at sulfur is preferred. This mechanistic distinction is mirrored in the classification of enzymes that catalyze Sulfate ester hydrolysis: arylsulfatases (ASs) catalyze S–O cleavage in Sulfate sugars and arylSulfates, and alkyl sulfatases break the C–O bond of alkyl Sulfates. Sinorhizobium meliloti choline sulfatase (SmCS) efficiently catalyzes the hydrolysis of alkyl Sulfate choline-O-Sulfate (kcat/KM = 4.8 × 103 s− 1 M− 1) as well as arylSulfate 4-nitrophenyl Sulfate (kcat/KM = 12 s− 1 M− 1). Its 2.8-A resolution X-ray structure shows a buried, largely hydrophobic active site in which a conserved glutamate (Glu386) plays a role in recognition of the quaternary ammonium group of the choline substrate. SmCS structurally resembles members of the alkaline phosphatase superfamily, being most closely related to dimeric ASs and tetrameric phosphonate monoester hydrolases. Although > 70% of the amino acids between protomers align structurally (RMSDs 1.79–1.99 A), the oligomeric structures show distinctly different packing and protomer–protomer interfaces. The latter also play an important role in active site formation. Mutagenesis of the conserved active site residues typical for ASs, H218O-labeling studies and the observation of catalytically promiscuous behavior toward phosphoesters confirm the close relation to alkaline phosphatase superfamily members and suggest that SmCS is an AS that catalyzes S–O cleavage in alkyl Sulfate esters with extreme catalytic proficiency.
