Mutant HMGR Enzyme Variants for Mevalonate-Pathway Squalene Production
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Solution Overview
Problem
Existing methods for producing squalene, an important intermediate in terpenoid synthesis, are inadequate, with production levels typically limited to about 5% by weight of dry fungal weight, and existing techniques for improving production conditions do not significantly exceed conventional levels.
Innovation Solution
The use of a hydroxymethylglutaryl CoA reductase enzyme with specific amino acid sequences, including variations at key positions in the Sα2 and Lα2 regions, to enhance the mevalonate pathway for increased squalene production.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional hydroxymethylglutaryl CoA reductase is used in the mevalonate pathway, then the pathway functions normally, but squalene production is limited to about 5% by weight of dry fungal weight
Solution Approach 1:
The patent applies parameter changes by modifying the amino acid sequence of hydroxymethylglutaryl CoA reductase at specific positions (Sα2 and Lα2 regions). This molecular-level parameter change results in enhanced enzyme activity and dramatically increased squalene production, resolving the contradiction between using conventional enzyme and achieving high productivity.
2Productivity
If existing techniques for improving production conditions are applied, then some optimization is achieved, but production levels do not significantly exceed conventional levels
Solution Approach 1:
Rather than进一步优化 production conditions, the patent achieves breakthrough by changing the fundamental parameter of the enzyme's amino acid sequence. This molecular modification provides a more effective solution than conventional process optimization, resolving the contradiction between productivity improvement and ease of manufacture.
3Productivity
If the mevalonate pathway is enhanced through conventional means, then terpenoid synthesis proceeds, but squalene production remains insufficient for large-scale application
Solution Approach 1:
The patent modifies the enzyme's amino acid sequence parameters to enhance the mevalonate pathway's squalene production capability. This molecular-level parameter change provides reliable and consistent high-level production, resolving the contradiction between production quantity and reliability for large-scale application.
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
This approach significantly increases the production of squalene, achieving levels comparable to or exceeding previous methods, demonstrating enhanced terpenoid compound synthesis efficiency.
Implementation Method 1
The use of a hydroxymethylglutaryl CoA reductase enzyme with specific amino acid sequences, including variations at key positions in the Sα2 and Lα2 regions, to enhance the mevalonate pathway for increased squalene production.
Data Source
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AI summary
The object of the present invention is to efficiently produce useful terpenoid compounds, and specifically, to provide a method for preparing squalene, which is an important intermediate of terpenoid. The object can be solved by a hydroxymethylglutaryl CoAreductase (HMGR) comprising:(a) an amino acid other than alanine (A) at the 10th position in an Sα2 amino acid sequence of HMGR, (b) an amino acid other than proline (P) at the 1 st position from the carboxyl terminal in the DKK region of the HMG-CoA binding site of HMGR, (c) an amino acid other than alanine (A) at the 1st position in an Lα2 amino acid sequence of HMGR, and (d) an amino acid other than glutamic acid (E) at the 6th position in an Lα2 amino acid sequence of HMGR of the present invention.