Development of a Polymerase for Genotyping

This case study outlines one of our life science enzyme development projects involving the scale-up and manufacture of a bespoke DNA polymerase enzyme and co-development of a suitable activity assay. DNA polymerase enzymes are used in DNA amplification, their function is to synthesise DNA molecules by assembling one nucleotide at a time. In industry, DNA polymerases have a wide range of application areas, from forensics to food contamination testing.

OVERVIEW: Our customer was manufacturing their DNA polymerase in-house at a small scale. Successful product launches lead to a rapidly expanding business where the increasing enzyme demand was expected to exceed their in-house capacity. Biocatalysts further developed and scaled-up the process to enable the manufacture of commercial quantities. A vital part of the development process included the co-development of an activity assay to allow for reliable quantification of enzyme activity to be used at both Biocatalysts’ and our customer’s site.

CUSTOMER CHALLENGE: The manufacture of a bespoke DNA polymerase complete with an activity assay for enzyme activity quantification.

APPROACH: The development of the manufacturing process involved optimising both the fermentation protocol and multiple down-stream processing (DSP; purification) steps to reach the highly specific quality specification. The development of this process can be broadly defined into three main stages:

1. Small-scale process development
Biocatalysts constructed an expression strain by transforming the customer’s plasmid into Biocatalysts’ E. coli strain. Expression trials were conducted to indicate the most suitable fermentation conditions to be used. A series of lab scale fermentations were then performed together with optimisation of small-scale DSP. This process provided test samples for the customer to confirm enzyme performance in their applications.

2. Assay development
Throughout the process development, we worked closely with our customer to improve and develop an existing activity assay. The addition of Biocatalysts’ experience in enzyme assay development resulted in a robust and reliable method of enzyme activity quantification that is used at both Biocatalysts’ and the customer’s sites.

3. Scale up of the whole process
The final step in this project was the scale-up of the fermentation and DSP to provide commercial quantities to the customer, meeting the pre-defined specification. From the beginning of the project Biocatalysts always had commercial scale manufacture in mind, therefore, all manufacturing steps were purposely selected and tailored to allow for a seamless scale up and to allow the flexibility to quickly produce at an even larger scale if required in the future. The successful project led to a long-term enzyme supply agreement with the customer.