Lactobacillus casei Promoter Surface Expression Vector

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Solution Overview

Problem

Current methods for expressing foreign proteins on bacterial surfaces face limitations due to insufficient expression levels and instability, particularly when using inducible promoters, and existing surface anchoring motifs do not adequately support the stable display of larger proteins.

Innovation Solution

A Lactobacillus casei-derived galactose mutarotase gene promoter and a poly-gamma-glutamate synthetase complex gene from a Bacillus sp. strain are used to create a surface expression vector that stably anchors target proteins on the microbial surface, enhancing expression levels and stability.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If conventional inducible promoters are used for expressing foreign proteins on bacterial surfaces, then expression can be controlled, but expression levels are insufficient and stability is poor

Engineering Contradiction:

Improveexpression stabilityVSAvoidexpression level

Core Design Contradiction:

ReliabilityVSProductivity

Solution Approach 1:

The patent changes the promoter parameter from conventional inducible promoters to a novel promoter derived from Lactobacillus casei. This promoter exhibits both high expression levels and high stability without requiring induction, fundamentally changing the expression parameters to simultaneously achieve high productivity and reliability

Inventive Principle:

Principle #35Parameter changes

2Stability of the object's composition

If existing surface anchoring motifs are used, then proteins can be displayed on the surface, but larger proteins cannot be stably displayed

Engineering Contradiction:

Improveprotein display stabilityVSAvoidprotein size accommodation

Core Design Contradiction:

Stability of the object's compositionVSAdaptability or versatility

Solution Approach 1:

The patent introduces an outer membrane protein (OmpA) from Bacillus subtilis as an intermediary anchoring motif. This OmpA protein serves as a mediator that can stably anchor various foreign proteins including larger proteins on the bacterial surface, overcoming the limitations of existing anchoring motifs while maintaining stable display

Inventive Principle:

Principle #24Intermediary (Mediator)

Data Source

PatentUS12180469B2Surface expression vector for constitutive high-expression using promoter of galactose mutarotase gene derived from <i>Lactobacillus casei</i>, and use thereof

Publication Date: 2024.12.31 BIOLEADERS CORP

AI summary

Provided is a galactose mutarotase gene promoter derived from Lactobacillus casei and the use thereof, and more particularly, to a Lactobacillus casei-derived galactose mutarotase gene promoter having the nucleotide sequence of SEQ ID NO: 1, an expression vector containing the promoter, and a microorganism transformed with the expression vector. A microorganism transformed with an expression vector containing the promoter may effectively express a target protein on the cell surface, and thus is useful as a vaccine vehicle or the like. Moreover, provided is a surface expression vector having pgsA, which is a gene encoding poly-gamma-glutamate synthetase, and a method of expressing a target protein on the microbial surface using the vector. The vector containing foreign genes inserted therein is transformed into a microorganism and allows a foreign protein is to be stably expressed on the surface of the microorganism.