Protein Science

HitGen Protein Expression Platform

HitGen’s protein expression platform encompasses a comprehensive suite of systems, including E. coli, insect cells, mammalian cells, and yeast (Figure 1).

We specialize in producing proteins tailored to specific project needs and client requirements, starting with rational construct design. Our services include a variety of tag options, functional labeling (e.g., biotinylation), and defined post-translational modifications. Most target gene constructs are meticulously designed based on literature data, specific downstream application needs, and extensive experience.

All purified proteins undergo stringent quality control, including SDS-PAGE and SEC-HPLC to ensure over 90% purity and homogeneity. Validation is achieved through LC/MS and functional/binding assays (where available or feasible to develop). Proteins are typically delivered in multi-milligram quantities. For challenging targets, we conduct focused small-scale screens to identify optimal boundaries and fusion tags, enhancing production success rates.

In response to the growing demand for Targeted Protein Degradation (TPD) technology, HitGen has successfully produced over 50 different E3 ubiquitin ligases, a key focus for TPD platforms. To date, we have obtained dozens of high-quality E3 subfamily substrate recognition proteins and their complexes (Table 1).

Additionally, HitGen routinely produces a variety of tool enzymes for specific downstream applications, including high-quality T4 DNA ligase in gram quantities to support internal and external requirements for DNA Encoded Library (DEL) construction, and dozens of amino acid synthetases etc. Over the past decade, our in-house expressed proteins have covered more than 40 different target classes (Figure 2).

High-quality proteins are essential for accelerating key processes in drug discovery. HitGen’s protein production capabilities seamlessly integrate with other services, including:

• X-ray crystallography for SBDD and FBDD programs

• In-vitro assay development

• DEL-based screening for hit identification

Our “crystallography-grade” recombinant protein production is available as a stand-alone service or combined with crystal structure determination. We also offer comprehensive crystallization services, including:

• Initial crystallization screening using high-throughput nanoliter dispensing robots (Mosquito™), automated crystal imaging (UVEX-ps256, Jansi), and in-house X-ray diffraction (Bruker D8 VENTURE with METAJET). (Figure 3)

• Multi-dimensional optimization for optimal crystal conditions.

• X-ray diffraction and data collection (monthly schedule at SSRF) followed by crystal structure determination (Figure 4).

Figure 1

Table 1

Figure 2

Figure 3

Figure 4