Halophilic PHB Depolymerase Mutations for High-Salt Recycling
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Solution Overview
Problem
Existing PHA depolymerases are not conducive to industrial processes due to insufficient thermodynamic and thermal stability, kinetic speed, and inability to function in environments contaminated with human waste, limiting the use of biopolymers in consumer products and industrial recycling.
Innovation Solution
Development of halophilic and thermophilic PHB depolymerases with single-site mutations that enhance solubility, stability, and kinetic characteristics, enabling their use in high-salt, high-temperature industrial processes for simultaneous degradation and decontamination of biopolymers, particularly in post-consumer personal care products.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If natural PHA depolymerases are used for industrial processing, then biodegradation of PHA polymers can be achieved, but the enzymes lack sufficient thermodynamic and thermal stability, kinetic speed, and ability to function in contaminated environments
Solution Approach 1:
The patent applies parameter changes by modifying the enzyme's amino acid sequence through site-directed mutagenesis. Specific residues are mutated to alter the enzyme's physical and chemical properties, including thermal stability, solubility, and catalytic activity. This allows the enzyme to maintain functionality under industrial processing conditions while improving productivity.
Solution Approach 2:
The patent creates modified copies of natural PHA depolymerases by introducing specific mutations into the enzyme gene. These engineered variants retain the core catalytic function of natural enzymes but possess improved properties for industrial applications, effectively copying and enhancing the original enzyme's capabilities.
2Stability of the object's composition
If natural PHA depolymerases are used, then PHA polymer degradation can occur, but the enzymes are not soluble or stable in high-salt, high-temperature industrial environments
Solution Approach 1:
The patent modifies enzyme parameters by changing amino acid residues that affect solubility and thermal stability. Mutations are introduced to enhance the enzyme's ability to remain soluble and stable in high-salt, high-temperature conditions, thereby improving environmental adaptability for industrial processing.
Solution Approach 2:
The patent applies local quality changes by targeting specific regions of the enzyme molecule for mutation. Rather than modifying the entire enzyme, specific amino acid residues are altered to confer local improvements in solubility and stability without affecting the overall catalytic function.
3Productivity
If natural PHA depolymerases are used for rapid degradation, then substrate conversion can be achieved, but the enzymes lack kinetic speed required for minimal processing time
Solution Approach 1:
The patent changes kinetic parameters of the enzyme by mutating residues involved in substrate binding and catalysis. These modifications increase the catalytic rate constant (kcat) and decrease the Michaelis constant (Km), resulting in faster substrate conversion and reduced processing time for industrial applications.
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 PHB depolymerases exhibit improved solubility, stability, and kinetic performance, allowing for efficient degradation and decontamination of biopolymers in industrial settings, even in the presence of contaminants like feces and urine, thereby supporting sustainable recycling and waste reduction.
Implementation Method 1
catalysts for industrial degradation of PHA polymers... PHB depolymerases... degradation of the polymer into monomers
Data Source
AI summary
The present invention relates to a method for treatment of poly hydroxy alkanoate (PHA) containing post-consumer product, the method comprising contacting a post-consumer product with a polypeptide that can catalyze degradation of the PHA, the contact taking place at a temperature at least 40° C. and in the presence of salt at a concentration of 1 M or greater. In a specific embodiment, the polypeptide is a wild-type PHA depolymerase expressed by a halophilic microorganism or a modified PHA depolymerase that includes one or more single-site mutations as compared to the wild-type PHA depolymerase. In another specific embodiment, the polypeptide comprising a modified poly hydroxy butyrate (PHB) depolymerase comprising one or more single-site mutations as compared to SEQ ID NO: 1, and the modified PHB depolymerase having a solubility of 10 mg/L or greater. The present invention also relates to a host cell transformed to express a polypeptide that catalyzes degradation of a PHA in the presence of salt at a concentration of 1 M or greater, wherein the host cell is selected from an E. coli cell or a halophilic microorganism.