scholarly journals Subcellular localization and properties of mouse adrenal C19-steroid 5β-reductase

1975 ◽  
Vol 147 (1) ◽  
pp. 165-174 ◽  
Author(s):  
W Collins ◽  
E H Cameron
Keyword(s):  
Substrate Concentration ◽  
Soluble Fraction ◽  
Product Concentration ◽  
High Substrate Concentration ◽  
Ph Optimum ◽  
Metabolizing Enzymes ◽  
Small Activity ◽  
Substrate Concentrations ◽  
Hydroxy Steroid ◽  
Steroid Dehydrogenase

The localization and some characteristics of mouse adrenal C19-steroid 5 β-reductase were determined by the incubation of subcellular fractions of mouse adrenal tissue with [7 α-3H]androst-4-ene-3,17-dione. This enzyme was present only in the soluble fraction and was NADPH-dependent, although a small activity in the presence of NADH was also detected. The soluble fraction also contained 3α-, 3β- and a small amount of 17 β-hydroxy steroid dehydrogenase. These and other steroid-metabolizing enzymes present in the remaining subcelluar fractions are also described briefly. To measure 5 β-androstane-3,17-dione production by the mouse adrenal soluble fraction, all 5 β products first had to be oxidized to 5 β-androstane-3,17-dione, and the recovery of radio-activity between the substrate androst-4-ene-3,17-dione and product 5 β-androstane-3,17-dione of 96.1 +/-3.2% validated this technique. C19-steroid 5 β-reductase has a pH optimum of 6.5 and at low substrate concentrations the Km and Vmax. for 5 β reduction of [7 α-3H]androst-4-ene-ene-3,17-dione was 2.22 times 10(-6) ±0.48 times 10(-6) M and 450+/- 53 pmol/min per mg of protein respectively. At high substrate concentration, inhibition of the reaction occurred, which was shown to be due to increasing product concentration.

scholarly journals Metabolism of androst-4-ene-3,17-dione by subcellular fractions of rat adrenal tissue with particular reference to microsomal C19-steroid 5α-reductase

1973 ◽  
Vol 132 (2) ◽  
pp. 283-291 ◽  
Author(s):  
Paul V. Maynard ◽  
Euan H. D. Cameron
Keyword(s):  
Cytochrome C ◽  
Microsomal Fraction ◽  
Soluble Fraction ◽  
Subcellular Fractions ◽  
Dehydrogenase System ◽  
Separate System ◽  
Hydroxy Steroid ◽  
Microsomal Pellet ◽  
Steroid Dehydrogenase

The location and some characteristics of rat adrenal C19-steroid 5α-reductase were investigated by using [7α-3H]androst-4-ene-3,17-dione and [7α-3H]testosterone as substrates. The enzymes system was shown to be NADPH-dependent and associated with the microsomal fraction. In addition, some evidence was also obtained for the existence of a separate NADH-dependent system in the soluble fraction. Further investigation of androst-4-ene-3,17-dione metabolism by subcellular fractions indicated the presence of NADH-dependent 3α- and 3β-hydroxy steroid dehydrogenase systems in the microsomal pellet. This pellet also appeared to contain an NADH-dependent 17β-hydroxy steroid dehydrogenase system, and a similar though separate system was detected in the cytosol. Malate (20mm) effectively inhibited the microsomal C19-steroid 5α-reductase, which showed similar values for Km and Vmax. when either androst-4-ene-3,17-dione or testosterone was used as substrate. Cytochrome c was added to all incubation mixtures used for the determination of these values to inhibit the formation of metabolites other than 5α-androstane-3,17-dione and 5α-dihydrotestosterone (17β-hydroxy-5α-androstan-3-one) respectively. It was also found that corticosterone did not inhibit the 5α-reduction of androst-4-ene-3,17-dione under these conditions, indicating that separate enzymes exist for the 5α-reduction of C19- and C21-steroids in the rat adrenal.

scholarly journals Isolation and characterization of 17β-hydroxy steroid dehydrogenase from human erythrocytes

1972 ◽  
Vol 127 (4) ◽  
pp. 649-659 ◽  
Author(s):  
E. Mulder ◽  
G. J. M. Lamers-Stahlhofen ◽  
H. J. Van Der Molen
Keyword(s):  
Dehydrogenase Activity ◽  
Gel Filtration ◽  
Enzymic Activity ◽  
Hydroxyl Group ◽  
Unit Weight ◽  
Ph Optimum ◽  
Ammonium Sulphate Precipitation ◽  
Isolation And Characterization ◽  
Hydroxy Steroid ◽  
Steroid Dehydrogenase

1. The 17β-hydroxy steroid dehydrogenase was solubilized during haemolysis of erythrocytes and was isolated from the membrane-free haemolysate. Membrane preparations isolated in different ways did not contain 17β-hydroxy steroid dehydrogenase activity. The 17β-hydroxy steroid dehydrogenase activity in the haemolysate was concentrated by repeated ammonium sulphate precipitation and gel filtration on Sephadex G-150. The 17β-hydroxy steroid dehydrogenase activity of the purified preparation per unit weight of protein was 350–3000 times higher than the activity of the crude erythrocyte haemolysate. The 20α-hydroxy steroid dehydrogenase activity was lost during this purification procedure. 2. The 17β-hydroxy steroid dehydrogenase was NADP-dependent and had a pH optimum for conversion of testosterone between 8.5 and 10. For the molecular weight of the enzyme a value of 64000 was calculated from Sephadex chromatography results. 3. p-Chloromercuribenzoate inhibited the enzymic activity. The oxidative activity of the enzyme for the 17β-hydroxyl group was only partly inhibited when a large excess of 17-oxo steroids was added. The catalysing activity of the enzyme was influenced by the NADP+/NADPH ratio. The oxidation of the 17β-hydroxyl group in the presence of NADP+ proceeded faster than the reduction of the 17-oxo group with NADPH. When both reduced and oxidized cofactors were present the oxidation of the 17β-hydroxyl group was inhibited to a considerable extent. 4. The enzyme had a broad substrate specificity and not only catalysed the conversion of androstanes with a 17β-hydroxyl group, or 17-oxo group, but also the conversion oestradiol⇆oestrone. In addition the steroid conjugates dehydroepiandrosterone sulphate and oestrone sulphate were also converted. There were no indications that more than one 17β-hydroxy steroid dehydrogenase was present in the partially purified preparation.

Multiplication of a Klebsiella pneumoniae Strain in Water at Low Concentrations of Substrates

1988 ◽  
Vol 20 (11-12) ◽  
pp. 117-123 ◽  
Author(s):  
D. van der Kooij ◽  
W. A. M. Hijnen
Keyword(s):  
Amino Acids ◽  
Water Treatment ◽  
Klebsiella Pneumoniae ◽  
Substrate Concentration ◽  
Treatment System ◽  
Water Treatment System ◽  
Low Concentrations ◽  
Ks Values ◽  
Substrate Concentrations

A K.pneumoniae strain, isolated from a water treatment system, was tested in growth measurements for its ability to multiply at substrate concentrations of a few micrograms per liter. The organism multiplied on mixtures of carbohydrates and amino acids at a substrate concentration of 1 µg of C of each compound per liter. Tests with individual compounds revealed that especially carbohydrates were utilized at low concentrations. The Ks values obtained for maltose and maltopentaose were 53 µg of C/l and 114 µg of C per liter, respectively. The significance of the growth of K.pneumoniae at low substrate concentrations is discussed.

A metabolic stability determination of Tetrahydrothiazolopyridine derivative a selective 11β-hydroxy steroid dehydrogenase type 1 (11β-hsd1) inhibitor

2017 ◽  
Vol 8 (3) ◽  
Author(s):  
MURUGESH KANDASAMY ◽  
MUHAMMED SALIHIN ◽  
MALLIKARJUNA RAO PICHIKA ◽  
SLAVKO KOMARNYTSKY ◽  
THIRUMURUGAN RATHINASABAPATHY
Keyword(s):  
Metabolic Stability ◽  
Hydroxy Steroid ◽  
Steroid Dehydrogenase

Notizen: Effect of Magnetic Field on Ascorbic Acid Oxidase Activity, I

1986 ◽  
Vol 41 (3) ◽  
pp. 355-358 ◽  
Author(s):  
V. S. Ghole ◽  
P. S. Damle ◽  
W. H.-P. Thiemann
Keyword(s):  
Magnetic Field ◽  
Ascorbic Acid ◽  
Homogeneous Magnetic Field ◽  
Acid Oxidase ◽  
High Substrate Concentration ◽  
Ascorbic Acid Oxidase ◽  
Rate Of Reaction ◽  
Substrate Concentrations ◽  
Burk Plot

A homogeneous magnetic field of 1.1 T strength exhibits a significant influence on the activity of the enzyme ascorbic acid oxidase in vitro. A Lineweaver-Burk plot of the reaction shows the typical pattern of a mixed-type inhibition, i.e. a larger rate of reaction at low substrate concentrations and a smaller rate of reaction at high substrate concentration than that of the control without magnetic field applied.

scholarly journals The determination of binding parameters when the total and free substrate concentrations are not approximately equal

1979 ◽  
Vol 179 (3) ◽  
pp. 697-700 ◽  
Author(s):  
N Gains
Keyword(s):  
Substrate Concentration ◽  
Graphical Method ◽  
Enzyme Concentration ◽  
Binding Parameters ◽  
Free Concentration ◽  
Equilibrium Binding ◽  
Substrate Concentrations

By using a standard graphical method values of Km and V may be found that are independent of the conditions and assumptions that the total substrate concentration approximates to its free concentration and that Km is much larger than the enzyme concentration. The procedure is also applicable to the determination of equilibrium binding parameters of a ligand to a macromolecule.

DIFFICULTIES IN THE DIAGNOSIS OF THE ADRENOGENITAL SYNDROME IN INFANCY

PEDIATRICS ◽  
1972 ◽  
Vol 49 (2) ◽  
pp. 198-205
Author(s):  
C. H. Shackleton ◽  
F. L. Mitchell ◽  
J. W. Farquhar
Keyword(s):  
Hydroxylase Deficiency ◽  
Adrenogenital Syndrome ◽  
Steroid Excretion ◽  
21 Hydroxylase Deficiency ◽  
Hydroxy Steroid ◽  
Steroid Dehydrogenase

Pregnanetriol was not excreted by an infant (7 days old) who was later shown to have a defect in steroid 21-hydroxylase. However, the excretion of this compound increased during the following days (1.2 mg on the thirteenth day of life). A high excretion of 3β-hydroxy-Δ steroids was the most noticeable abnormality in steroid excretion noted on the seventh day of life (e.g., 3β, 16α-dihydroxy-5-pregnen-20-one, 15 mg; 3β, 21-dihydroxy-5-pregnen-20-one, 1.4 mg and 3β, 16α-dihydroxy-5-androsten-17-one, 7.4 mg). This high 3β-hydroxy-Δ steroid excretion results in difficulties in distinguishing a defect in 3β-hydroxy steroid dehydrogenase from a 21-hydroxylase deficiency. At the age of 14 months the principal steroids excreted were those predominant in other cases of 21-hydroxylase deficiency, viz. pregnanetriol and 5β-pregnane-3α, 17α, 20α-triol-11-one (11-oxo-pregnanetriol).

scholarly journals Crystal structure of Arabidopsis thaliana neutral invertase 2

2020 ◽  
Vol 76 (3) ◽  
pp. 152-157
Author(s):  
Łukasz P. Tarkowski ◽  
Vicky G. Tsirkone ◽  
Evgenii M. Osipov ◽  
Steven Beelen ◽  
Willem Lammens ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Glycoside Hydrolase Family ◽  
Plant Cell Walls ◽  
Higher Plant ◽  
Ph Optimum ◽  
Metabolizing Enzymes ◽  
Neutral Invertase ◽  
Sucrose Metabolizing Enzymes ◽  
Life On Earth ◽  
Hydrolase Family

The metabolism of sucrose is of crucial importance for life on Earth. In plants, enzymes called invertases split sucrose into glucose and fructose, contributing to the regulation of metabolic fluxes. Invertases differ in their localization and pH optimum. Acidic invertases present in plant cell walls and vacuoles belong to glycoside hydrolase family 32 (GH32) and have an all-β structure. In contrast, neutral invertases are located in the cytosol and organelles such as chloroplasts and mitochondria. These poorly understood enzymes are classified into a separate GH100 family. Recent crystal structures of the closely related neutral invertases InvA and InvB from the cyanobacterium Anabaena revealed a predominantly α-helical fold with unique features compared with other sucrose-metabolizing enzymes. Here, a neutral invertase (AtNIN2) from the model plant Arabidopsis thaliana was heterologously expressed, purified and crystallized. As a result, the first neutral invertase structure from a higher plant has been obtained at 3.4 Å resolution. The hexameric AtNIN2 structure is highly similar to that of InvA, pointing to high evolutionary conservation of neutral invertases.

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