| Research Abstract |
Phospholipids are classified into glycerophospholipids and sphingophospholipids. Phospholipase AィイD22ィエD2 (PLAィイD22ィエD2) catalyzes the hydrolysis of the ester linkage at the sn-2 position of glycerophospholipids and responsibles for the metabolism of unsaturated fatty acids. Meanwhile, sphingomyelinase is the general term of the enzyme which catalyzes the hydrolysis of the phosphoester linkage of sphingomyelin to produce ceramide, which has been known as a lipid second messenger in mammalian cell membranes.
We found that 3-methoxycarbonyl-2, 4, 6-trienal (A) strongly inactivated phospholipase AィイD22ィエD2 which was isolated from bovine pancreas. The inactivation mechanism of PLAィイD22ィエD2 by the compound A was elucidated by means of the stereocontrolled synthesis of the derivatives of A, structure activity relationship, amino acid analysis of the complex with PLAィイD22ィエD2, and model reactions with amines. Thus, the compound A was found to selectively modify Lys-56 which is included in the
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interfacial recognition site of bovine pancreatic PLAィイD22ィエD2, by the MALDI-TOF-Mass peptide mapping analyses of the complex.
We also synthesized the new cyclic amide analog as the second-generation inhibitor having an oxazolidinone ring, because we had previously synthesized the oxazolidinone phospholipid analog which competitively acted on the catalytic site of PLAィイD22ィエD2.
On the other hand, in the study of sphingomyelinase, we achieved the efficient synthesis of (D)-erythro-sphigomyelin, and found that (D)-erythro stereochemistry is essentially important as the substrate for sphingomyelinase and the double bond in the backbone skeleton was not essential for the hydrolysis by B. cereus sphingomyelinase. Furthermore, we established the stereocontrolled efficient synthesis of sphingomyelin methylene and ethylene analogs, which are the first substrate-analog inhibitors, and they actually inhibited the hydrolytic ability of the enzyme. These analog inhibitors might be useful for the elucidation of the sphingomyelinase hydrolytic mechanism. Less
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