Structural Basis for Substrate Specificity in Human Monomeric Carbonyl Reductases

Abstract

Carbonyl reduction constitutes a phase I reaction for many xenobiotics and is carried out in mammals mainly by members of two protein families, namely aldo-keto reductases and short-chain dehydrogenases/reductases. In addition to their capacity to reduce xenobiotics, several of the enzymes act on endogenous compounds such as steroids or eicosanoids. One of the major carbonyl reducing enzymes found in humans is carbonyl reductase 1 (CBR1) with a very broad substrate spectrum. A paralog, carbonyl reductase 3 (CBR3) has about 70% sequence identity and has not been sufficiently characterized to date. Screening of a focused xenobiotic compound library revealed that CBR3 has narrower substrate specificity and acts on several orthoquinones, as well as isatin or the anticancer drug oracin. To further investigate structure-activity relationships between these enzymes we crystallized CBR3, performed substrate docking, site-directed mutagenesis and compared its kinetic features to CBR1. Despite high sequence similarities, the active sites differ in shape and surface properties. The data reveal that the differences in substrate specificity are largely due to a short segment of a substrate binding loop comprising critical residues Trp229/Pro230, Ala235/Asp236 as well as part of the active site formed by Met141/Gln142 in CBR1 and CBR3, respectively. The data suggest a minor role in xenobiotic metabolism for CBR3.Enhanced version This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.

Carbonyl Reduction of Mequindox by Chicken and Porcine Cytosol and Cloned Carbonyl Reductase 1

“…Likewise, the cCBR1 showed 6.97-fold higher k cat /K m than pCBR1 to MEQ (Table 3). The difference in catalytic activity toward MEN was similar to the result reported by other groups Pilka et al, 2009). They analyzed the substrate-binding site of human CBR1 and CBR3 by site-directed mutagenesis using isatin and 9,10-phenanthrenequinone as model substrates and demonstrated the importance of methionine 142 and tryptophan 230 in binding with the substrates.…”

Genetic variation in the carbonyl reductase 3 gene confers risk of type 2 diabetes and insulin resistance: a potential regulator of adipogenesis

“…However, we did not observe significant association between rs2066826 and type 2 diabetes in this study. The human CBR3 gene, a paralog of CBR1, encodes for a ketoreductase that converts the C-9 keto (carbonyl) group of PGE 2 to a hydroxyl group, leading to the formation of PGF2 α [29,30]. CBR1 is the major carbonyl reducing enzyme in humans, with a very broad substrate spectrum including steroids, eicosanoids, and xenobiotic compounds.…”

“…CBR1 is the major carbonyl reducing enzyme in humans, with a very broad substrate spectrum including steroids, eicosanoids, and xenobiotic compounds. CBR3 has about 70% sequence identity with CBR1 with a much narrower substrate spectrum [29,30]. PGE 2 has been shown to be the major prostaglandin produced in adipose tissue with potent adipogenic and anti-lipolytic effects through binding to the EP3 prostanoid receptors.…”

Curcumin is a tight-binding inhibitor of the most efficient human daunorubicin reductase – Carbonyl reductase 1

“…The presence of other reductases not inhibited by curcumin may explain the fact that despite the low K i -value for CBR1, residual activity was 40% even in the presence of 50 lM curcumin. C-13 carbonyl reduction of DAUN and DOX is mediated by several members of the SDR and AKR superfamilies, namely CBR1, CBR3, CBR4, AKR1A1, AKR1B1, AKR1B10, AKR1C1, AKR1C2, AKR1C3, AKR1C4 and AKR7A2 [36,[38][39][40][41], although reports concerning the role of CBR3 [41] and AKR1C2 in anthracycline metabolism are inconsistent [40,42].…”