Purification of oestrogen synthetase by high-performance liquid chromatography

Muto, Norio; Tan, Liat

doi:10.1016/s0021-9673(01)87439-2

Cited by 35 publications

(12 citation statements)

References 16 publications

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“…4 (Yoshimoto & Guengerich 2014 (Pasanen & Pelkonen 1981, Mendelson et al 1985, Muto & Tan 1985, Hall et al 1987, Kellis & Vickery 1987, Osawa et al 1987. These studies provided definitive evidence that aromatization involved a single enzyme, and not multiple enzymes as originally thought.…”

Section: Biosynthesis Of Oestrogens: Aromatase and Its Mechanismmentioning

confidence: 81%

“…In the 1980s, Y Osawa, P Hall, L Vickery, F Bellino, N Muto, M Pasonen, O Pelkonen, E Simpson, and C Mendelson purified the human aromatase cytochrome P450 protein from human placental microsomes and demonstrated conversion of androstenedione to oestrone with purified enzyme (Pasanen & Pelkonen 1981, Mendelson et al 1985, Muto & Tan 1985, Hall et al 1987, Kellis & Vickery 1987, Osawa et al 1987. These studies provided definitive evidence that aromatization involved a single enzyme, and not multiple enzymes as originally thought.…”

Section: Purificationmentioning

confidence: 89%

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Celebrating 75 years of oestradiol

Simpson

Santen

2015

Journal of Molecular Endocrinology

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Oestrogens exert important effects on the reproductive as well as many other organ systems in both men and women. The history of the discovery of oestrogens, the mechanisms of their synthesis, and their therapeutic applications are very important components of the fabric of endocrinology. These aspects provide the rationale for highlighting several key components of this story. Two investigators, Edward Doisy and Alfred Butenandt, purified and crystalized oestrone nearly simultaneously in 1929, and Doisy later discovered oestriol and oestradiol. Butenandt won the Nobel Prize for this work and Doisy's had to await his purification of vitamin K. Early investigators quickly recognized that oestrogens must be synthesized from androgens and later investigators called this process aromatization. The aromatase enzyme was then characterized, its mechanism determined, and its structure identified after successful crystallization. With the development of knock-out methodology, the precise effects of oestrogen in males and females were defined and clinical syndromes of deficiency and excess described. Their discovery ultimately led to the development of oral contraceptives, treatment of menopausal symptoms, therapies for breast cancer, and induction of fertility, among others. The history of the use of oestrogens for postmenopausal women to relieve symptoms has been characterized by cyclic periods of enthusiasm and concern. The individuals involved in these studies, the innovative thinking required, and the detailed understanding made possible by evolving biologic and molecular techniques provide many lessons for current endocrinologists. Discovery of oestrogenThe concept that substances secreted by one organ could then be transported internally to another via the blood stream is the underpinning of the field of endocrinology. The 'father of physiology', Claude Bernard, articulated the concept of internal secretion by describing how the liver produces glucose, which then travels to and is utilized by other tissues in the body. This concept underlies studies on the ovaries, which became a focus of investigation later in the century. In 1896, Emil Knauer, a 29-year-old Austrian, excised the ovaries of adult rabbits and then re-implanted pieces into different locations in the same animals (Watkins 2007 Doisy then enlisted a nurse in the obstetrics clinic to supply each pregnant patient with a 1 gal jug and ask for her urine. This provided the needed material but was fraught with difficulties. Suspected to be a bootlegger because of the large jugs of yellow material in his car, one of Doisy's drivers was nearly arrested until he suggested using the sniff test with the response 'My god, it is urine'. Doisy, then being recruited to St Louis University, continued his efforts to purify the active substance. Each extract was tested with a bioassay and its effect on uterine stimulation. Urine, the material of choice for study, was initially extracted with a very large countercurrent apparatus (Fig. 1). Costing the exorbitan...

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Section: Biosynthesis Of Oestrogens: Aromatase and Its Mechanismmentioning

confidence: 81%

Section: Purificationmentioning

confidence: 89%

Celebrating 75 years of oestradiol

Simpson

Santen

2015

Journal of Molecular Endocrinology

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“…Then, enzyme solution was dialyzed at 4 o C in distilled water and 50 mM K-acetate / 5mM K-phosphate buffer (pH 7.0) for 2 h with two changes of distilled water and buffer respectively (11). During all procedures, the temperature was kept at + 4 o C (11,17,18).…”

Section: Methodsmentioning

confidence: 99%

Purification and characterization of glucose 6-phosphate dehydrogenase from Lake Van fish (Chalcalburnus tarichii pallas, 1811) liver

Türkoğlu

Altun

Çiftçi

2006

J. Physiol. Biochem.

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Glucose 6-phosphate dehydrogenase (D-glucose 6-phosphate: NADP+ oxidoreductase, EC 1.1.1.49; G6PD) was purified from Lake Van fish (Chalcalburnus tarichii pallas, 1811) liver, using a simple and rapid method, and some characteristics of the enzyme were investigated. The purification procedure was composed of two steps: homogenate preparation and 2', 5'-ADP Sepharose 4B affinity gel chromatography, which took 7-8 hours. Thanks to the two consecutive procedures, the enzyme, having specific activity of 38 EU/mg protein, was purified with a yield of 44.39% and 1310 fold. In order to control the enzyme purification SDS polyacrylamide gel electrophoresis (SDS-PAGE) was done. SDS polyacrylamide gel electrophoresis showed a single band for enzyme. Optimal pH, stable pH, optimal temperature, Km and, Vmax values for NADP+ and glucose 6-phosphate (G6P) were also determined for the enzyme. In addition, molecular weight and subunit molecular weights were found by sodium dodecyl sulfate polyacrilamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography respectively.

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“…The protein was determined in the resultant solution. During all procedures, the temperature was kept at +4°C (Morelli et al, 1978;Muto and Tan, 1985;Ninfali et al, 1990).…”

Section: Methodsmentioning

confidence: 99%

Purification and characterization of glucose 6-phosphate dehydrogenase from rainbow trout (oncorhynchus mykiss) erythrocytes

Çiftçi¹,

Çiltaş²,

Erdoğan³

2004

Vet. Med. - Czech

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Glucose 6-phosphate dehydrogenase (D-glucose 6-phosphate: NADP + oxidoreductase, EC 1.1.1.49; G6PD) from rainbow trout (Oncorhynchus mykiss) erythrocytes was purified, using a simple and rapid method, and some characteristics of the enzyme were investigated. The purification procedure consisted of three steps: haemolysate preparation, ammonium sulphate precipitation, and 2',5'-ADP Sepharose 4B affinity gel chromatography, which took one working day. Thanks to the three consecutive procedures, the enzyme, having the specific activity of 14.51 EU/mg proteins, was purified with a yield of 70.40% and 1 271.19-fold. In order to control the purification of the enzyme SDS polyacrylamide gel electrophoresis was carried out. SDS polyacrylamide gel electrophoresis showed a single band for the enzyme. Optimal pH, stable pH, optimal temperature, molecular weight, and K M and V max values for NADP + and glucose 6-phosphate (G6-P) were also determined for the enzyme. In addition, the effect of NADPH on the enzyme was investigated and K i value and the type of inhibition were determined by means of Lineweaver-Burk graph obtained for NADPH.

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Purification of oestrogen synthetase by high-performance liquid chromatography

Cited by 35 publications

References 16 publications

Celebrating 75 years of oestradiol

Celebrating 75 years of oestradiol

Purification and characterization of glucose 6-phosphate dehydrogenase from Lake Van fish (Chalcalburnus tarichii pallas, 1811) liver

Purification and characterization of glucose 6-phosphate dehydrogenase from rainbow trout (oncorhynchus mykiss) erythrocytes

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