LysECD7 Variants for Spore Surface-Displayed Microbial Lysis
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Solution Overview
Problem
The rise of antibiotic-resistant microbes poses a significant threat to human health, and there is a need for alternative antimicrobial therapies, particularly antimicrobial peptides (AMPs) that can effectively target a broad range of pathogens, including both gram-negative and gram-positive bacteria, and address biofilm formation on implants.
Innovation Solution
Development of LysECD7 variants with specific amino acid substitutions and fusion proteins, such as LysECD7-SMAP, which are expressed on spores or associated with microbes, providing enhanced antimicrobial activity and specificity through surface display or fusion with crust proteins like CotY, and utilizing modified spores with non-germinating genomes for targeted microbial inhibition.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional antibiotics are used to treat bacterial infections, then bacterial growth is inhibited, but antibiotic resistance develops reducing treatment effectiveness
Solution Approach 1:
The patent modifies the amino acid sequence of the LysECD7 lysin protein to create variants with altered parameters including enhanced catalytic activity, improved stability under different conditions, and modified target specificity. These parameter changes enable the protein to maintain effectiveness against resistant bacterial strains by optimizing its enzymatic mechanism of action rather than relying on conventional antibiotic binding mechanisms.
Solution Approach 2:
The patent creates fusion proteins that combine LysECD7 lysin with other functional domains or peptides (such as cell wall binding domains or antimicrobial peptides) to produce composite antimicrobial agents. These composite structures integrate multiple functions including enhanced bacterial binding, improved stability, and amplified lysis activity, thereby overcoming resistance mechanisms that single-function antibiotics cannot address.
2Productivity
If LysECD7 variants with specific amino acid substitutions are developed, then microbial lysis efficiency is improved, but protein structure complexity increases
Solution Approach 1:
The patent introduces specific amino acid substitutions at targeted positions within the LysECD7 protein structure, particularly in regions that contact the bacterial cell wall or catalytic domains. These localized modifications enhance lysis efficiency at specific functional sites without requiring comprehensive restructuring of the entire protein, thereby improving productivity while limiting complexity increases to only essential regions.
Solution Approach 2:
The patent designs LysECD7 variants with enhanced conformational flexibility or induced fit mechanisms that allow the protein to dynamically adapt to different bacterial cell wall structures. This dynamic behavior enables the protein to maintain high lysis efficiency across diverse targets through controlled structural adjustments rather than requiring complex static structures, optimizing function while managing structural complexity.
3Adaptability or versatility
If LysECD7-SMAP fusion protein is created to broaden antimicrobial spectrum, then activity against both gram-negative and gram-positive bacteria is achieved, but molecular weight increases
Solution Approach 1:
The patent fuses the LysECD7 lysin domain with the SMAP-29 antimicrobial peptide domain to create a single fusion protein that combines the cell wall lytic activity of LysECD7 with the membrane-disrupting activity of SMAP-29. This merging approach broadens the antimicrobial spectrum to cover both gram-negative and gram-positive bacteria through a single molecular entity, achieving versatility while avoiding the need for separate administration of multiple agents.
Solution Approach 2:
The LysECD7-SMAP fusion protein is designed to perform multiple functions: LysECD7 provides endolytic activity against peptidoglycan in both gram-negative and gram-positive bacteria, while SMAP-29 contributes membrane-active antimicrobial properties. This multi-functional design creates a universal antimicrobial agent that can target different bacterial types and mechanisms simultaneously, achieving broad spectrum activity through a single optimized protein construct.
Data Source
AI summary
The present disclosure relates to a group of Antimicrobial peptides (referred to as “LysECD7 variants” or “LysECD7 variant proteins”) that are based on the wild-type enzyme LysECD7 but with specific amino acid substitutions, poly nucleotides and vectors encoding the same, peptide fragments thereof, cells or spores expressing the poly nucleotide or vector, and methods of using and making the same. In addition, the present disclosure relates to spore-associated or microbe-associated LysECD7 variants. The association of LysECD7 variants with spores or microbes can include surface display by covalent or noncovalent association or by mixing. The present disclosure is further directed to fragments or subsequences of LysECD7 variant nucleic acids comprising hybridizable portions of the LysECD7 variant sequence which have use. e.g. in nucleic acid-based assays.