Substrate Specificity in Short-Chain Phospholipid Analogs at the Active Site of Human Synovial Phospholipase A2
| Autor: | T N, Wheeler, S G, Blanchard, R C, Andrews, F, Fang, Y, Gray-Nunez, C O, Harris, M H, Lambert, M M, Mehrotra, D J, Parks, J A, Ray |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
Models
Molecular chemistry.chemical_classification Binding Sites biology Stereochemistry Substituent Phospholipid Active site Phospholipases A Enzyme assay Substrate Specificity Phospholipases A2 Structure-Activity Relationship chemistry.chemical_compound Phospholipase A2 Enzyme chemistry Enzymatic hydrolysis Critical micelle concentration Synovial Fluid Drug Discovery biology.protein Humans Molecular Medicine Phospholipids |
| Zdroj: | Journal of Medicinal Chemistry. 37:4118-4129 |
| ISSN: | 1520-4804 0022-2623 |
| DOI: | 10.1021/jm00050a009 |
| Popis: | The substrate specificity at the active site of recombinant human synovial fluid phospholipase A2 (hs-PLA2) was investigated by the preparation of a series of short-chain phospholipid analogs and measurement of their enzymatic hydrolysis at concentrations well below the critical micelle concentration. Substrates used in the study included 1,2-dihexanoylglycerophospholipids, 1,2-bis(alkanoylthio)glycerophospholipids, and 1-O-alkyl-2-(alkanoylthio)phospholipids. Turnover was observed for only a few of the 1,2-dihexanoylglycerophospholipids, and the rate of hydrolysis was very low, near the limit of detection of the assay. In contrast, selected 2-(alkanoylthio)-glycerophospholipids were hydrolyzed by hs-PLA2 at much higher rates at concentrations well below their critical micelle concentration (cmc). Thus, the 1,2-bis(hexanoylthio)glycerophosphatidylmethanol exhibits a k(cat)/K(M) = 1800 L mol-1 s-1. Over the calculated log P (cLogP) range of 3-9, cLogP and log(k(cat)/K(M) were linearly related for compounds with straight-chain sn-1 and sn-2 substituents. At comparable cLogP's, the sn-1 ethers and thioesters were hydrolyzed at comparable rates. A negative charge in the phosphate head group was required for enzyme activity. Unsaturation, aromaticity, and branching in the sn-2 substituent reduce turnover dramatically. The same structural modifications in the sn-1 substituent have less effect on turnover. Certain of these substrates, e.g., 1,2-bis(hexanoylthio)glycerophosphatidylmethanol, may be useful in assaying for active site inhibitors of PLA2. The structure--activity relationships established here for substrates should serve as a reference for the structure--activity relationships of substrate-based inhibitors. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|