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Tryptase from human pulmonary mast cells. Purification and characterization
The Journal of Biological Chemistry
|November 25, 1981
Summary
This study successfully purified tryptase, a key enzyme from human mast cells, revealing its tetrameric structure and active sites. This enzyme is present at high levels in mast cells, suggesting a significant role in tissue degranulation.
Area of Science:
- Biochemistry
- Immunology
Background:
- Mast cells are crucial immune cells containing potent proteases.
- Tryptase is the primary neutral protease found in human mast cell secretory granules.
Purpose of the Study:
- To purify and characterize tryptase from human pulmonary mast cells.
- To determine the enzyme's structure, molecular weight, and activity.
Main Methods:
- Sequential chromatography (Dowex 1-X2, DEAE-Sephadex, heparin-agarose) for purification.
- Polyacrylamide gel electrophoresis (native and denaturing) for structural analysis.
- Gel filtration chromatography for molecular weight determination.
Main Results:
- Tryptase was purified to homogeneity, showing a single band on native PAGE.
- The holoenzyme has a molecular weight of 144,000 Da, composed of four subunits (37,000 and 35,000 Da).
- All subunits demonstrated active site capacity, and the enzyme exhibited high specific activity for tosyl-L-arginine methyl ester.
Conclusions:
- Human pulmonary mast cells contain significantly higher levels of tryptase compared to other leukocytes.
- Purified tryptase is a tetrameric enzyme with potential for high enzymatic activity.
- Mast cell degranulation can release substantial amounts of tryptase into tissues.