| 1. |
- Benkestock, Kurt, et al.
(författare)
-
Automated Nano-Electrospray Mass Spectrometry for Protein-Ligand Screening by Noncovalent Interaction Applied to Human H-FABP and A-FABP
- 2003
-
Ingår i: Journal of Biomolecular Screening. - : Elsevier BV. - 1087-0571 .- 1552-454X. ; 8:3, s. 247-256
-
Tidskriftsartikel (refereegranskat)abstract
- A method for ligand screening by automated nano-electrospray ionization mass spectrometry (nano-ESI/MS) is described. The core of the system consisted of a chip-based platform for automated sample delivery from a 96-well plate and subsequent analysis based on noncovalent interactions. Human fatty acid binding protein, H-FABP (heart) and A-FABP (adipose), with small potential ligands was analyzed. The technique has been compared with a previously reported method based on nuclear magnetic resonance (NMR), and excellent correlation with the found hits was obtained. In the current MS screening method, the cycle time per sample was 1.1 min, which is approximately 50 times faster than NMR for single compounds and approximately 5 times faster for compound mixtures. High reproducibility was achieved, and the protein consumption was in the range of 88 to 100 picomoles per sample. Furthermore, a novel protocol for preparation of A-FABP without the natural ligand is presented. The described screening approach is suitable for ligand screening very early in the drug discovery process before conventional high-throughput screens (HTS) are developed and/or used as a secondary screening for ligands identified by HTS.
|
|
| 2. |
- Hämäläinen, Markku D., et al.
(författare)
-
Characterization of a set of HIV-1 protease inhibitors using binding kinetics data from a biosensor-based screen
- 2000
-
Ingår i: Journal of Biomolecular Screening. - : Elsevier BV. - 1087-0571 .- 1552-454X. ; 5:5, s. 353-359
-
Tidskriftsartikel (refereegranskat)abstract
- The interaction between 290 structurally diverse human immunodeficiency virus type 1 (HIV-1) protease inhibitors and the immobilized enzyme was analyzed with an optical biosensor, Although only a single concentration of inhibitor was used, information about the kinetics of the interaction could be obtained by extracting binding signals at discrete time points. The statistical correlation between the biosensor binding data, inhibition of enzyme activity (K-i), and viral replication (EC50) revealed that the association and dissociation rates for the interaction could be resolved and that they were characteristic for the compounds. The most potent inhibitors, with respect to K-i and EC50 values, including the clinically used drugs, all exhibited fast association and slow dissociation rates. Selective or partially selective binders for HIV-1 protease could be distinguished from compounds that showed a general protein-binding tendency by using three reference target proteins. This biosensor-based direct binding assay revealed a capacity to efficiently provide high-resolution information on the interaction kinetics and specificity of the interaction of a set of compounds with several targets simultaneously.
|
|
| 3. |
- Lindholm, Petra, et al.
(författare)
-
Selective cytotoxicity evaluation in anticancer drug screening of fractionated plant extracts
- 2002
-
Ingår i: Journal of Biomolecular Screening. - : Elsevier BV. - 1087-0571 .- 1552-454X. ; 7:4, s. 333-340
-
Tidskriftsartikel (refereegranskat)abstract
- Chosen to reflect biodiversity in a phylogenetic sense, 100 fractionated plant extracts were screened in vitro for cytotoxicity following extraction and fractionation (polypeptide isolation). Of these 100 extracts, 30 were selected and then characterized preliminarily for antitumor potency and mode of action by testing them on two cell lines and primary cultures of human tumor cells. On the basis of cytotoxicity potency, 10 of the extracts were further characterized for anticancer activity in 10 human tumor cell lines. This final testing resulted in seven potential lead plants with superior evidence of antitumor potential: Colchicum autumnale L. (Colchicaceae), Digitalis lanata Ehrh. and Digitalis purpurea L. (Plantaginaceae), Helleborus cyclophyllus Boiss. (Ranunculaceae), Menyanthes trifoliata L. (Menyanthaceae), and Viola arvensis Murr. and Viola patrinii Ging. (Violaceae). Within a database of antitumor compounds, the activity profiles of the extracts from these seven plants were compared, by correlation analysis, with those of more than 100 other compounds, including 39 standard drugs from different classes of cytotoxic mechanisms. The activity profiles of six of these candidates were uncorrelated with those of the standard drugs, possibly indicating new pathways of drug-mediated cell death.
|
|
