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Literature summary for 1.14.14.23 extracted from

  • Tempel, W.; Grabovec, I.; MacKenzie, F.; Dichenko, Y.V.; Usanov, S.A.; Gilep, A.A.; Park, H.W.; Strushkevich, N.
    Structural characterization of human cholesterol 7alpha-hydroxylase (2014), J. Lipid Res., 55, 1925-1932.
    View publication on PubMedView publication on EuropePMC

Crystallization (Commentary)

Crystallization (Comment) Organism
wild-type CYP7A1 in the ligand-free state, structure-based mutation T104L in the B' helix, corresponding to the nonpolar residue of CYP7B1, is used to obtain crystals of complexes with cholest-4-en-3 -one and with cholesterol oxidation product 7-ketocholesterol, hanging drop vapor diffusion method, mixing 0f 0.001 ml of protein solution with 0.001 ml of reservoir solution containing 0.1 M sodium chloride, 0.1 M trisodium citrate, pH 5.5, and 20% PEG 400, 18°C, to obtain a complex with cholest-4-en-3-one, a T104L mutant is used, and the crystals are grown in conditions containing 0.1 M MES, pH 6.0, and 18% PEG 550 monomethylether and soaked with 20% glycerol. For the T104L mutant-7KCh complex, crystals are grown in 0.1 M sodium chloride, 0.1 M tri-sodium citrate, pH 5.6, and 20% PEG 400, room temperature, X-ray diffraction structure determination and analysis at 1.90-2.75 A resolution Homo sapiens

Protein Variants

Protein Variants Comment Organism
T104L the mutant is similar to the wild-type in ligand binding and catalytic properties and readily crystallizes with substrates. The structure of the T104L mutant in complex with cholest-4-en-3-one explicitly identifies key residues involved in 7alpha-hydroxylation Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
cholesterol + [reduced NADPH-hemoprotein reductase] + O2 Homo sapiens
-
7alpha-hydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
?

Organism

Organism UniProt Comment Textmining
Homo sapiens P22680
-
-

Source Tissue

Source Tissue Comment Organism Textmining
liver
-
Homo sapiens
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
25-hydroxycholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
Homo sapiens ?
-
?
27-hydroxycholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
Homo sapiens ?
-
?
cholest-4-en-3-one + [reduced NADPH-hemoprotein reductase] + O2
-
Homo sapiens ?
-
?
cholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
Homo sapiens 7alpha-hydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
?
cholesterol + [reduced NADPH-hemoprotein reductase] + O2 stereospecific hydroxylation of cholesterol Homo sapiens 7alpha-hydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
?
additional information substrate spcecificity and recognition structure, CYP7A1 can bind and hydroxylate cholesterol and cholest-4-en-3-one with high efficiency as well as oxysterols, overview. Cholest-4-en-3-one bound parallel to the heme with the alpha-side of the steroid nucleus facing the heme plane at a distance of about 3.5 A, substrate binding strutcure, detailed overview Homo sapiens ?
-
?

Synonyms

Synonyms Comment Organism
cholesterol 7alpha-hydroxylase
-
Homo sapiens
CYP7A1
-
Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Homo sapiens

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.003
-
cholest-4-en-3-one pH 7.2, 37°C, wild-type enzyme Homo sapiens
0.008
-
cholesterol pH 7.2, 37°C, wild-type enzyme Homo sapiens
0.015
-
25-hydroxycholesterol pH 7.2, 37°C, wild-type enzyme Homo sapiens
0.023
-
cholesterol pH 7.2, 37°C, T104L mutant enzyme Homo sapiens
0.0685
-
27-Hydroxycholesterol pH 7.2, 37°C, wild-type enzyme Homo sapiens
0.117
-
cholest-4-en-3-one pH 7.2, 37°C, T104L mutant enzyme Homo sapiens
0.117
-
27-Hydroxycholesterol pH 7.2, 37°C, T104L mutant enzyme Homo sapiens

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.2
-
assay at Homo sapiens

Cofactor

Cofactor Comment Organism Structure
heme
-
Homo sapiens
NADPH
-
Homo sapiens

General Information

General Information Comment Organism
metabolism hepatic conversion to bile acids is a major elimination route for cholesterol in mammals. CYP7A1 catalyzes the first and rate-limiting step in classic bile acid biosynthesis, converting cholesterol to 7alpha-hydroxycholesterol. Hydroxylation of the ring system of cholesterol in a regio- and stereospecific manner with further oxidation and shortening of the side chain produces water-soluble bile acids with powerful detergent properties to emulsify dietary lipids. Bile acids also serve as signaling molecules that bind to G-protein-coupled receptors and nuclear hormone receptors that regulate lipid, glucose, and energy metabolism Homo sapiens
additional information identification of a motif of residues that promote cholest-4-en-3-one binding parallel to the heme, thus positioning the C7 atom for hydroxylation. Orientation of cholesterol in the different leaflets of the lipid bilayer can be recognized for further binding by CYP7A1 alone or in complex with protein partners Homo sapiens
physiological function hepatic conversion to bile acids is a major elimination route for cholesterol in mammals. CYP7A1 catalyzes the first and rate-limiting step in classic bile acid biosynthesis, converting cholesterol to 7alpha-hydroxycholesterol Homo sapiens