Recombinant Human Parathyroid Hormone (rH-PTH) Engineering Service

Recombinant Human Parathyroid Hormone (rH-PTH), particularly the bioactive fragment PTH (1-34) (Teriparatide), is a crucial biopharmaceutical used to treat osteoporosis by stimulating bone formation. The production faces a dilemma: inactivity in prokaryotic expression (E. coli) due to misfolding and lack of necessary post-translational modifications, and high cost in eukaryotic expression (CHO, yeast) due to complex media, slow growth, and expensive purification. The key is finding a way to make the cost-effective E. coli system produce a correctly folded, active product.

CD Biosynsis offers a synthetic biology service focused on achieving high-quality production of rH-PTH using E. coli. Our core strategy involves modification of the Escherichia coli fusion expression system where PTH is fused to a highly soluble protein tag (e.g., SUMO, MBP) to promote high expression and prevent aggregation into inactive inclusion bodies. This is followed by the optimization of the protein refolding process using specialized buffers, additives, and controlled environmental conditions to maximize the yield of monomeric, correctly folded, and active rH-PTH (1-34) after cleavage of the fusion tag. This integrated approach aims to deliver a high-yield, high-activity, and cost-effective bioproduction route.

Pain Points

Establishing an economically viable rH-PTH production route with E. coli faces these key technical challenges:

• Inactivity in Prokaryotic Expression: PTH is highly sensitive to folding errors. When expressed in E. coli without assistance, it often forms inactive inclusion bodies or misfolds rapidly, resulting in little to no biological activity.
• High Cost in Eukaryotic Expression: While mammalian or yeast systems can ensure better folding, they require expensive cell culture media, low-productivity fermentation , and complex purification, making the final product costly.
• Refolding Yield: Even when inclusion bodies are successfully solubilized, the subsequent in vitro refolding yield is often very low (typically under 10%), making the entire process inefficient.
• Proteolytic Degradation: The PTH (1-34) fragment is a small peptide highly susceptible to degradation by host-cell proteases or during downstream processing.

A cost-effective solution must maximize the yield of correctly folded, active rH-PTH from the high-productivity E. coli host.

Solutions

CD Biosynsis utilizes advanced protein engineering and bioprocess optimization to produce active rH-PTH in E. coli:

Modification of E. coli Fusion Expression System

           

We fuse the PTH sequence to a highly soluble fusion tag (SUMO, MBP) linked by a specific protease cleavage site to maximize soluble expression and ease purification.

Optimization of Protein Refolding Process

We use high-throughput screening of refolding conditions (detergents, denaturants, redox buffers) to identify the optimal environment for efficient PTH monomer formation.

Codon and mRNA Optimization

We optimize the PTH gene sequence for high expression in E. coli and engineer the mRNA structure to enhance translational speed and stability .

Co-expression of Folding Chaperones

We test co-expression of bacterial chaperones (e.g., GroEL}/\text{GroES) to assist in correct in vivo folding and potentially increase the soluble fraction of the fusion protein.

This systematic approach is focused on overcoming the folding hurdle inherent in producing small, complex peptides in E. coli.

Advantages

Our rH-PTH engineering service is dedicated to pursuing the following production goals:

High Refolding Yield and Purity

Optimized refolding protocols significantly increase the conversion rate to active, monomeric rH-PTH .

Cost-Effective E. coli Platform

Using high-productivity E. coli drastically reduces upstream fermentation costs compared to slow, expensive eukaryotic systems.

High Specific Activity

Validation ensures the final product is correctly folded, resulting in maximal biological activity for therapeutic use. [Image of Cost Reduction Icon]

Simplified Inclusion Body Handling

Fusion tag use often leads to consistent, high-purity inclusion bodies , simplifying the initial recovery steps.

PTH (1-34) Specific Production

The system is specifically tailored to produce the therapeutically relevant PTH (1-34) fragment efficiently, avoiding full-length PTH complexity.

We provide a specialized platform aimed at maximizing the yield and minimizing the purification cost of active rH-PTH biomanufacturing.

Process

Our rH-PTH engineering service follows a rigorous, multi-stage research workflow:

• Fusion Vector Construction: Design the expression vector with an optimized fusion tag (e.g., MBP, SUMO) and a highly specific, efficient cleavage site between the tag and PTH (1-34).
• Inclusion Body Preparation: Optimize E. coli induction conditions for maximum accumulation of fusion protein into high-purity inclusion bodies.
• Refolding Condition Screening: Use automated screening systems to test a wide array of buffers, pH levels, reducing agents, and additives to identify the optimal high-yield refolding cocktail .
• Cleavage and Purification: Optimize the enzymatic or chemical cleavage step to release PTH (1-34), followed by high-resolution chromatography (RP-HPLC) to isolate the monomeric, active product.
• Quality and Activity Validation: Characterize the final product using mass spectrometry, CD spectroscopy, and bioactivity assays (e.g., adenylyl cyclase stimulation) to ensure correct structure and function.
• Result Report Output: Compile a detailed Experimental Report including vector maps, refolding protocols, and final titer/activity data , supporting CMC documentation.

Technical communication is maintained throughout the process, focusing on timely feedback regarding fusion protein yield and final refolding efficiency.

Explore the potential for a cost-effective, high-quality rH-PTH supply. CD Biosynsis provides customized protein and process solutions:

• Detailed Refolding Optimization and Activity Report , demonstrating the improved conversion rate to active PTH.
• Consultation on downstream RP-HPLC purification strategies for achieving pharmaceutical-grade purity.
• Experimental reports include complete raw data on inclusion body yield and final active product purity , essential for regulatory submissions.