Article

Insights in 17β-HSD1 Enzyme Kinetics and Ligand Binding by Dynamic Motion Investigation

Pharmaceutical and Medicinal Chemistry, Saarland University, Saarbrücken, Germany.
PLoS ONE (Impact Factor: 3.53). 08/2010; 5(8):e12026. DOI: 10.1371/journal.pone.0012026
Source: PubMed

ABSTRACT Bisubstrate enzymes, such as 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1), exist in solution as an ensemble of conformations. 17beta-HSD1 catalyzes the last step of the biosynthesis of estradiol and, thus, it is a potentially attractive target for breast cancer treatment.
To elucidate the conformational transitions of its catalytic cycle, a structural analysis of all available crystal structures was performed and representative conformations were assigned to each step of the putative kinetic mechanism. To cover most of the conformational space, all-atom molecular dynamic simulations were performed using the four crystallographic structures best describing apoform, opened, occluded and closed state of 17beta-HSD1 as starting structures. With three of them, binary and ternary complexes were built with NADPH and NADPH-estrone, respectively, while two were investigated as apoform. Free energy calculations were performed in order to judge more accurately which of the MD complexes describes a specific kinetic step.
Remarkably, the analysis of the eight long range trajectories resulting from this multi-trajectory/-complex approach revealed an essential role played by the backbone and side chain motions, especially of the betaF alphaG'-loop, in cofactor and substrate binding. Thus, a selected-fit mechanism is suggested for 17beta-HSD1, where ligand-binding induced concerted motions of the FG-segment and the C-terminal part guide the enzyme along its preferred catalytic pathway. Overall, we could assign different enzyme conformations to the five steps of the random bi-bi kinetic cycle of 17beta-HSD1 and we could postulate a preferred pathway for it. This study lays the basis for more-targeted biochemical studies on 17beta-HSD1, as well as for the design of specific inhibitors of this enzyme. Moreover, it provides a useful guideline for other enzymes, also characterized by a rigid core and a flexible region directing their catalysis.

Download full-text

Full-text

Available from: Matthias Negri, Jun 27, 2015
1 Follower
 · 
118 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: In order to become and remain competitive, organizations across America are moving away from traditional hierarchical management toward a network of small work teams. Recent surveys suggest that as many as 80% of American businesses now have one or more work teams in place. Such teams, dedicated to the continuous improvement of their piece of the business, have the potential of becoming the primary unit of performance in these new corporate systems. Many self-directed, problem solving and cross-functional teams are producing dramatic results in quality, productivity and speed. Far too many, however, are failing to realize their potential. Some never seem to get off the ground while others plateau or even drop off in performance after a promising start. For many organizations, keeping the momentum going is even more of a challenge than getting them started in the first place. Based on the author's experience in training and developing high performance teams, this paper identifies the three major factors that contribute to team performance. Additionally, it describes fifteen characteristics that must be addressed in order for work teams to reach their performance potential. Using these characteristics as a checklist can be helpful when establishing new work teams. But they may be even more useful when attempting to determine why existing work teams have leveled off or even decreased in performance
    Engineering Management Conference, 1994. 'Management in Transition: Engineering a Changing World', Proceedings of the 1994 IEEE International; 11/1994
  • Value in Health 11/2008; 11(6). DOI:10.1016/S1098-3015(10)66810-6 · 2.89 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Estradiol (E2), the most important estrogen in humans, is involved in the initiation and progression of estrogen-dependent diseases such as breast cancer and endometriosis. Its local production in the target cell is regulated by 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1), which catalyzes E2-formation by reduction of the weak estrogen estrone (E1). Because the enzyme is expressed in the diseased tissues, inhibition of 17β-HSD1 is considered as a promising therapy for the treatment of estrogen-dependent diseases. For the development of novel inhibitors, a structure- and ligand-based design strategy was applied, resulting in bicyclic substituted hydroxyphenylmethanones. In vitro testing revealed high inhibitory potencies toward human placental 17β-HSD1. Compounds were further evaluated with regard to selectivity (17β-HSD2, estrogen receptors ERα and ERβ), intracellular activity (T47D cells), and metabolic stability. The most promising compounds, 14 and 15, showed IC(50) values in the low nanomolar range in the cell-free and cellular assays (8-27 nM), more than 30-fold selectivity toward 17β-HSD2 and no affinity toward the ERs. The data obtained make these inhibitors interesting candidates for further preclinical evaluation.
    Journal of Medicinal Chemistry 10/2010; 53(22):8176-86. DOI:10.1021/jm101073q · 5.48 Impact Factor