Recombinant Hemoglobins with Engineered Self-Purification
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Solution Overview
Problem
Existing recombinant hemoglobins have low oxygen carrying capacity and are prepared using laborious and costly methods with high impurity levels, including protoporphyrin-IX (PPIX), which impairs their oxygen carrier function, and require a heat treatment step that reduces yield and necessitates a costly CO removal process.
Innovation Solution
Development of recombinant hemoglobins with improved oxygen-carrying capacity, comprising a di-alpha chain and two beta chains with specific sequence homologies, produced using a method that eliminates the need for heat treatment and CO removal, resulting in high yield and purity with reduced PPIX levels.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If heat treatment step is used to remove impurities, then purity of recombinant hemoglobin is improved, but yield is reduced and cost increases
Solution Approach 1:
The patent applies preliminary action by engineering the hemoglobin protein sequence in advance to include temperature-sensitive instability regions. This allows the protein to naturally precipitate impurities like PPIX during controlled cooling after expression, eliminating the need for separate heat treatment steps and preserving yield while achieving high purity.
Solution Approach 2:
The recombinant hemoglobin is designed to self-purify through its engineered temperature-sensitive properties. During controlled cooling, the hemoglobin selectively precipitates bound impurities such as PPIX, allowing the protein to perform its own purification function without external intervention, thereby maintaining high yield and purity simultaneously.
2Manufacturing precision
If heat treatment step is incorporated, then impurity removal is improved, but manufacturing cost increases
Solution Approach 1:
The patent applies preliminary action by engineering the hemoglobin protein sequence in advance to include temperature-sensitive instability regions. This allows the protein to naturally precipitate impurities like PPIX during controlled cooling after expression, eliminating the need for separate heat treatment steps and preserving yield while achieving high purity.
Solution Approach 2:
The recombinant hemoglobin is designed to self-purify through its engineered temperature-sensitive properties. During controlled cooling, the hemoglobin selectively precipitates bound impurities such as PPIX, allowing the protein to perform its own purification function without external intervention, thereby maintaining high yield and purity simultaneously.
3Stability of the object's composition
If CO is used to maintain stability during heat treatment, then stability is improved, but product purity worsens due to CO contamination
Solution Approach 1:
The patent applies parameter changes by eliminating the need for CO from the process. Instead of using CO to maintain stability during heat treatment, the invention uses controlled temperature changes (cooling) to achieve both stability and impurity removal, thereby avoiding CO contamination while maintaining product purity.
4Ease of manufacture
If conventional recombinant hemoglobin production is used, then production simplicity is improved, but oxygen carrying capacity worsens due to PPIX impurity
Solution Approach 1:
The patent applies preliminary action by engineering the hemoglobin protein sequence in advance to include temperature-sensitive instability regions. This allows the protein to naturally precipitate impurities like PPIX during controlled cooling after expression, eliminating the need for separate heat treatment steps and preserving yield while achieving high purity.
Solution Approach 2:
The recombinant hemoglobin is designed to self-purify through its engineered temperature-sensitive properties. During controlled cooling, the hemoglobin selectively precipitates bound impurities such as PPIX, allowing the protein to perform its own purification function without external intervention, thereby maintaining high yield and purity simultaneously.
Data Source
AI summary
The present disclosure relates to recombinant hemoglobins with improved oxygen carrying capacity and methods of preparation and use thereof. The present disclosure also relates to a pharmaceutical composition including the recombinant hemoglobin useful for treating oxygen deficiency related diseases or conditions, such as stroke, hemorrhagic shock, peripheral arterial disease (PAD), acute mountain sickness (AMS), cancer, and Parkinson's disease (PD).