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Titolo:
Kinetic alteration of a human dihydrodiol/3 alpha-hydroxysteroid dehydrogenase isoenzyme, AKR1C4, by replacement of histidine-216 with tyrosine or phenylalanine
Autore:
Ohta, T; Ishikura, S; Shintani, S; Usami, N; Hara, A;
Indirizzi:
Gifu Pharmaceut Univ, Biochem Lab, Gifu 5028585, Japan Gifu Pharmaceut Univ Gifu Japan 5028585 Biochem Lab, Gifu 5028585, Japan
Titolo Testata:
BIOCHEMICAL JOURNAL
, volume: 352, anno: 2000,
parte:, 3
pagine: 685 - 691
SICI:
0264-6021(200012)352:<685:KAOAHD>2.0.ZU;2-C
Fonte:
ISI
Lingua:
ENG
Soggetto:
3-ALPHA-HYDROXYSTEROID DIHYDRODIOL DEHYDROGENASE; HUMAN ALDOSE REDUCTASE; SITE-DIRECTED MUTAGENESIS; HUMAN LIVER; SUBSTRATE-SPECIFICITY; STEROID RECOGNITION; MOLECULAR-CLONING; PROSTAGLANDIN D-2; CRYSTAL-STRUCTURE; MESSENGER-RNA;
Keywords:
aldo-keto reductase; coenzyme binding; non-essential activator; substrate-binding cleft;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
39
Recensione:
Indirizzi per estratti:
Indirizzo: Hara, A Gifu Pharmaceut Univ, Biochem Lab, Gifu 5028585, Japan Gifu Pharmaceut Univ Gifu Japan 5028585 Lab, Gifu 5028585, Japan
Citazione:
T. Ohta et al., "Kinetic alteration of a human dihydrodiol/3 alpha-hydroxysteroid dehydrogenase isoenzyme, AKR1C4, by replacement of histidine-216 with tyrosine or phenylalanine", BIOCHEM J, 352, 2000, pp. 685-691

Abstract

Human dihydrodiol dehydrogenase with 3 alpha -hydroxysteroid dehydrogenaseactivity exists in four forms (AKR1C1-1C4) that belong to the aldo-keto reductase (AKR) family. Recent crystallographic studies on the other proteinsin this family have indicated a role for a tyrosine residue (correspondingto position 216 in these isoenzymes) in stacking the nicotinamide ring of the coenzyme. This tyrosine residue is conserved in most AKR family membersincluding AKR1C1-1C3, but is replaced with histidine in AKR1C4 and phenylalanine in some AKR members, In the present study we prepared mutant enzymesof AKR1C4 in which His-216 was replaced with tyrosine or phenylalanine. The two mutations decreased 3-fold the K-m for NADP(+) and differently influenced the K-m and k(cat) for substrates depending on their structures. The kinetic constants for bile acids with a 12 alpha -hydroxy group were decreased 1.5-7-fold and those for the other substrates were increased 1.3-9-fold. The mutation also yielded different changes in sensitivity to competitive inhibitors such as hexoestrol analogues, 17 beta -oestradiol, phenolphthalein and flufenamic acid and 3,5,3',5'-tetraiodothyropropionic acid analogues. Furthermore, the mutation decreased the stimulatory effects of the enzymeactivity by sulphobromophthalein, clofibric acid and thyroxine, which increased the K-m for the coenzyme and substrate of the mutant enzymes more highly than those of the wild-type enzyme. These results indicate the importance of this histidine residue in creating the cavity of the substrate-binding site of AKR1C4 through the orientation of the nicotinamide ring of the coenzyme, as well as its involvement in the conformational change by binding non-essential activators.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 10/07/20 alle ore 08:50:35