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Structure of an engineered, metal-actuated switch in trypsin
M E McGrath1, B L Haymore, N L Summers
1Department of Biochemistry and Biophysics, University of California, San Francisco 94143.
Biochemistry
|March 2, 1993
Summary
The copper complex of rat trypsin mutant R96H shows His57 reorienting to bind copper with His96. This specific active site reorganization causes metal-induced enzymatic inhibition.
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- Rat trypsin is a serine protease with a critical active site.
- Mutating Arg96 to His96 (trypsin R96H) engineers a potential metal-binding site.
- Metal ions can inhibit trypsin R96H activity.
Purpose of the Study:
- Determine the X-ray crystal structure of the copper complex of trypsin R96H.
- Ascertain the nature of the engineered metal-binding site.
- Understand the structural basis for metal-induced enzymatic inhibition.
Main Methods:
- X-ray crystallography was used to determine the structure of the copper-trypsin R96H complex.
- Chemical data analysis was employed to identify the copper ligand.
Main Results:
- His57 reorients from the active site to form a chelating metal-binding site with His96.
- Copper is bound by His57 and His96, with a third ligand, likely Tris.
- A water molecule occupies the active site, hydrogen-bonding with Asp102 and His57.
- The trypsin R96H structure is largely similar to native trypsin, except for active site rearrangements.
Conclusions:
- The engineered metal-binding site in trypsin R96H involves His57 and His96.
- Metal-induced inhibition of trypsin R96H is caused by a specific, reversible active site reorganization.