ImpE Protein Variants for High-Yield Microbial IMP Export

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Solution Overview

Problem

Existing methods for producing 5'-inosine monophosphate (IMP) through microbial fermentation face challenges in achieving high yield due to suboptimal enzyme properties and insufficient structural information, limiting effective enzyme application and IMP export efficiency.

Innovation Solution

Identification and utilization of protein variants, specifically ImpE1 and ImpE2, derived from Corynebacterium species, which enhance the activity of exporting IMP, along with the use of vectors and polynucleotides to express these proteins in microorganisms, thereby improving IMP production and export.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If rational design and site-directed mutagenesis of enzymes are applied to improve enzyme function, then enzyme activity can be enhanced, but the approach is disadvantageous because information on the structure of target enzymes is not sufficient or the structure-function correlation is not clear

Engineering Contradiction:

Improveenzyme functionVSAvoidstructure information

Core Design Contradiction:

ReliabilityVSLoss of information

Solution Approach 1:

The patent applies parameter changes by systematically varying amino acid residues at specific positions in the ImpE protein sequence. Through site-directed mutagenesis, individual residues were modified to alter protein function, ultimately identifying the critical G64D mutation that enhanced IMP export activity by 1.7-fold compared to the wild-type protein.

Inventive Principle:

Principle #35Parameter changes

2Reliability

If directed evolution method is used to improve enzyme activity through random mutagenesis and screening, then enzymes of desired traits can be obtained, but the process is time-consuming and requires screening large libraries of modified enzymes

Engineering Contradiction:

Improveenzyme activityVSAvoidscreening time

Core Design Contradiction:

ReliabilityVSLoss of time

Solution Approach 1:

The patent employed preliminary action by using bioinformatics analysis and sequence alignment to identify conserved regions and potential functional sites in the ImpE protein before conducting mutagenesis. This pre-analysis guided the selection of specific residues for mutation, reducing the need for extensive random screening and accelerating the discovery of functional improvements.

Inventive Principle:

Principle #10Preliminary action

3Productivity

If wild-type ImpE protein is used for IMP export, then the natural export function is maintained, but the export efficiency is insufficient to achieve high-yield IMP production

Engineering Contradiction:

ImproveIMP production yieldVSAvoidexport activity

Core Design Contradiction:

ProductivityVSReliability

Solution Approach 1:

The patent applied local quality by making a specific localized change at position 64 of the ImpE protein, where glycine was replaced with aspartic acid. This single localized mutation in the protein sequence significantly enhanced the export function, demonstrating that improving a specific local region can lead to overall functional enhancement without altering the entire protein structure.

Inventive Principle:

Principle #3Local quality

Data Source

PatentEP3674313B1"novel polypeptide and method of producing imp using the same

Publication Date: 2025.11.26 CJ CHEILJEDANG CORP

AI summary

The present disclosure relates to a novel protein variant having an activity of exporting 5'-inosine monophosphate, a microorganism comprising the protein variant, and a method for preparing 5'-inosine monophosphate using the microorganism.