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Structure of alpha-lytic protease complexed with its pro region
1Howard Hughes Medical Institute, University of California, San Francisco, 94143-0448, USA.
Nature Structural Biology
|November 10, 1998
Summary
The pro region (Pro) of alpha-lytic protease (alphaLP) acts as a folding catalyst, accelerating protein folding by a factor of 3 x 10^9. Structural analysis reveals how Pro stabilizes the transition state, overcoming alphaLP's significant folding barrier.
Area of Science:
- Biochemistry
- Structural Biology
- Protein Folding Dynamics
Background:
- Most proteins fold spontaneously, but alpha-lytic protease (alphaLP) exhibits an exceptionally slow folding rate (t(1/2) ≈ 2,000 years) due to a high folding barrier (30 kcal mol(-1)).
- AlphaLP is synthesized as a pro-enzyme, with its pro region (Pro) essential for facilitating proper folding.
- The Pro region acts as a potent folding catalyst, accelerating alphaLP folding by a factor of 3 x 10^9 when supplied separately.
Purpose of the Study:
- To elucidate the structural basis for the catalytic role of the Pro region in alphaLP folding.
- To understand the mechanism by which Pro overcomes the substantial folding barrier of alphaLP.
- To investigate the interaction between Pro and both the native and intermediate states of alphaLP.
Main Methods:
- X-ray crystallography was employed to determine the structures of the Pro region and the complex of Pro with native alphaLP.
- Analysis of protein interfaces, including hydration patterns, was performed to understand binding interactions.
- Functional assays were utilized to assess the catalytic activity of Pro on alphaLP folding.
Main Results:
- The crystal structure reveals a C-shaped Pro region that forms a large, complementary interface with the C-terminal beta-barrel domain of native alphaLP.
- Extensive hydration within the Pro-alphaLP interface explains tight binding to the native state and even tighter binding to the folding transition state.
- In the absence of Pro, alphaLP forms a stable molten globule-like intermediate, which rapidly folds to the native state upon Pro addition.
Conclusions:
- The Pro region's structure and its extensive, hydrated interface with alphaLP are key to its catalytic function.
- Pro stabilizes the folding transition state, thereby overcoming the significant kinetic barrier to alphaLP folding.
- A specific structural element within alphaLP is proposed as the primary cause of the folding barrier, with Pro providing a mechanism to circumvent it.