Protease-Resistant Beta-Hexosaminidase B Homodimer for Enzyme Replacement
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Solution Overview
Problem
Current enzyme replacement therapies for Tay-Sachs and Sandhoff diseases face challenges such as adverse reactions due to enzyme recognition as foreign matter, low stability in blood, and low cellular uptake, as well as protease degradation of the administered recombinant Hex A enzyme.
Innovation Solution
A modified β-subunit of β-hexosaminidase (ModB) is developed with specific amino acid substitutions to mimic the α-subunit's activity and resist protease degradation, forming a homodimer with enhanced stability and cellular uptake.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If wild-type recombinant Hex A is administered to treat Tay-Sachs or Sandhoff disease, then the enzyme can degrade GM2 gangliosides, but the enzyme is recognized as foreign matter and triggers adverse immune reactions
Solution Approach 1:
The patent applies parameter changes by modifying the amino acid sequence of the β-subunit through site-directed mutagenesis. Specifically, the active site residues are changed to mimic the α-subunit's active site configuration, creating a modified β-subunit that forms a homodimer with altered immunogenic properties while retaining enzymatic activity against GM2 gangliosides
Solution Approach 2:
The invention copies the functional characteristics of the α-subunit into the β-subunit by transferring the active site sequence. The modified β-subunit (ModB) is designed to have the same active site amino acid sequence as the α-subunit, allowing it to perform the same catalytic function while being structurally distinct enough to reduce immune recognition in α-subunit deficient patients
2Reliability
If wild-type recombinant Hex A is administered, then enzyme replacement therapy can be performed, but the enzyme shows low stability in blood and low cellular uptake
Solution Approach 1:
The patent modifies structural parameters of the enzyme by changing the amino acid sequence of the β-subunit. These sequence changes result in altered physical-chemical properties including enhanced stability in blood plasma and improved cellular uptake characteristics, while maintaining the essential catalytic function
3Reliability
If modified β-subunit (ModB) is administered to Sandhoff disease model mice, then GM2 gangliosides are degraded, but the enzyme is degraded by protease and symptoms are not improved
Solution Approach 1:
The patent applies parameter changes by modifying amino acid residues at positions known to be involved in protease recognition and cleavage. These sequence modifications create a protease-resistant modified β-subunit that maintains its structural integrity and enzymatic activity in the physiological environment, enabling sustained therapeutic effect
Solution Approach 2:
The invention applies preliminary anti-action by pre-modifying the enzyme structure to prevent protease degradation before the enzyme is administered. The amino acid substitutions are designed in advance to block protease recognition sites, thereby preventing the harmful degradation process before it can occur in the patient's body
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The modified ModB enzyme effectively degrades GM2 gangliosides with improved stability and reduced immune response, effectively treating Tay-Sachs and Sandhoff diseases by enhancing enzyme replacement therapy efficacy and reducing adverse reactions.
Implementation Method 1
the recombinant enzyme has the activity to degrade GM2 gangliosides
Data Source
AI summary
Provided is a modified .beta.-subunit of human .beta.-hexosaminidase which has the activity derived from the .alpha.-subunit of wild-type human .beta.-hexosaminidase and has the resistance to protease. A protein comprising an amino acid sequence having substitutions of the 312th to the 318th amino acids with glycine, serine, glutamic acid, proline, serine, glycine and threonine in order, respectively, in an amino acid sequence of a .beta.-subunit of wild-type human .beta.-hexosaminidase.