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Bioconversion of leukotriene D4 by lung dipeptidase
B J Campbell1, S F Baker, S D Shukla
1Department of Biochemistry, University of Missouri, Columbia 65212.
Biochimica Et Biophysica Acta
|January 16, 1990
Summary
Sheep lung dipeptidase, an enzyme involved in leukotriene metabolism, was successfully isolated and purified. This peptidase demonstrates significant activity and is inhibited by compounds like cilastatin, suggesting potential therapeutic applications.
Area of Science:
- Biochemistry
- Enzymology
- Membrane Protein Biochemistry
Background:
- Dipeptidases play crucial roles in cellular processes.
- Understanding the properties of lung dipeptidases is important for respiratory health.
- Membrane-bound enzymes often require specific methods for isolation and characterization.
Purpose of the Study:
- To isolate and purify sheep lung dipeptidase.
- To characterize its enzymatic activity, particularly its role in leukotriene metabolism.
- To investigate potential inhibitors of the enzyme.
Main Methods:
- Enzyme solubilization using phosphatidylinositol-specific phospholipase C.
- Purification via ammonium sulfate fractionation, affinity chromatography, and HPLC.
- Molecular weight estimation using HPLC.
- Enzymatic activity assays using glycyldehydrophenylalanine and leukotriene D4.
- Kinetic studies (Km, Vmax) and inhibitor screening.
Main Results:
- Sheep lung dipeptidase was effectively released from membrane preparations.
- Purified enzyme exhibited a molecular weight of approximately 105,000 Da.
- The enzyme demonstrated activity in converting leukotriene D4 to leukotriene E4.
- Kinetic parameters (Km, Vmax) were determined for leukotriene D4 hydrolysis.
- Cilastatin, bestatin, and chloracetyldehydrophenylalanine showed inhibitory effects, with cilastatin being the most potent.
Conclusions:
- Sheep lung dipeptidase is a membrane-anchored enzyme with significant activity in leukotriene metabolism.
- The enzyme's purification and characterization provide a basis for further functional studies.
- The identified inhibitors, particularly cilastatin, suggest potential therapeutic strategies targeting this enzyme in relevant physiological or pathological conditions.