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Visualization of Amyloid β Deposits in the Human Brain with Matrix-assisted Laser Desorption/Ionization Imaging Mass Spectrometry
Published on: March 7, 2019
Selection of mutant µplasmin for amyloid-β cleavage in vivo
Dongying Yang1, Wei Zhu2, Yingjie Wang3
1Shandong Provincial Key Laboratory of Biophysics, Shandong Key Laboratory in University of Functional Bioresource Utilization, School of Medicine and Nursing, Dezhou University, Daxuexi Road 566#, Dezhou, 253023, Shandong, China.
Abstract:
One of the main culprits of Alzheimer's disease (AD) is the formation of toxic amyloid-β (Aβ) peptide polymers and the aggregation of Aβ to form plaques in the brain. We have developed techniques to purify the catalytic domain of plasmin, micro-plasmin (µPlm), which can be used for an Aβ-clearance based AD therapy. However, in serum, µPlm is irreversibly inhibited by its principal inhibitor α2-antiplasmin (α2-AP). In this study, we engineered and selected mutant forms of µPlm that are both catalytically active and insensitive to α2-AP inhibition. We identified surface residues of μPlm that might interact and bind α2-AP, and used an alanine-scanning mutagenesis method to select residues having higher activity but lower α2-AP inhibition. Then we employed saturation mutagenesis for further optimize both properties. Modeled complex structure of µPlm/α2-AP shows that F587 is a critical contact residue, which can be used as a starting position for further investigation.
Insights
Researchers engineered micro-plasmin (µPlm) to resist inhibition by α2-antiplasmin (α2-AP), creating a potential Alzheimer's disease (AD) therapy targeting amyloid-β (Aβ) plaque clearance.
Area of Science:
- Biochemistry
- Neuroscience
- Enzyme Engineering
Background:
- Alzheimer's disease (AD) is characterized by amyloid-β (Aβ) plaque formation.
- Micro-plasmin (µPlm) shows potential for Aβ-clearance therapy.
- Serum inhibitor α2-antiplasmin (α2-AP) irreversibly inhibits µPlm.
Purpose of the Study:
- To engineer catalytically active and α2-AP-insensitive µPlm mutants.
- To develop an Aβ-clearance based Alzheimer's disease therapy.
Main Methods:
- Alanine-scanning mutagenesis to identify residues affecting µPlm/α2-AP interaction.
- Saturation mutagenesis to optimize µPlm activity and α2-AP resistance.
- Computational modeling of the µPlm/α2-AP complex structure.
Main Results:
- Identified surface residues critical for α2-AP binding.
- Selected µPlm mutants with enhanced activity and reduced α2-AP inhibition.
- F587 identified as a key contact residue in the µPlm/α2-AP complex.
Conclusions:
- Engineered µPlm variants offer a promising therapeutic strategy for Alzheimer's disease.
- Targeting the µPlm/α2-AP interaction can overcome inhibitory challenges.
- Further investigation of F587 is warranted for therapeutic development.
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