Incorporation of Unnatural Amino Acids into Proteins with e.coli expression systems

Background

The incorporation of unnatural amino acids (Uaas) is a breakthrough technology in the field of protein engineering. It greatly expands the structural and functional diversity of proteins and achieves precise modification of proteins by expanding the genetic code. This method not only enriches the functionality of proteins, but also provides a powerful new tool for studying protein structure, function and their interactions. Traditionally, proteins are composed of 20 standard amino acids, but by introducing UAAs, proteins can be endowed with new chemical properties and biological functions, such as fluorescent labeling, photocrosslinking and specific chemical modifications.

The E. coli expression system has become an ideal platform for the incorporation of unnatural amino acids due to its advantages such as simple operation, low cost and rapid growth. By modifying the translation machinery of E. coli, specific stop codons (usually the amber codon UAG) can be recoded to carry unnatural amino acids. This process requires two key components: an orthogonal aminoacyl-tRNA synthetase (aaRS) and the corresponding tRNA. This pair of orthogonal components can specifically recognize the target unnatural amino acid and accurately incorporate it into the specified site in the protein. Optimizing E. coli culture conditions, including temperature, nutrients, and induction timing, can increase the successful incorporation rate of UAAs and protein yield. These optimization measures are crucial to ensuring the quality of the final product. With the advancement of synthetic biology, more and more UAAs have been successfully applied to various types of proteins, making this technology show great potential in drug development, biosensor design, basic biological research, etc.

Our technology platform is characterized by high incorporation efficiency and high protein expression. By optimizing codon usage, regulatory element selection and culture conditions, we are able to provide customers with milligram to gram quantities of proteins containing unnatural amino acids. Whether it is used for structural biology research such as X-ray crystallography, NMR, cryo-electron microscopy, or for functional analysis and drug screening, our services can meet the high standards of our customers.

Service Procedure

Profacgen provide one-stop services from gene design to protein purification. Customers only need to provide the gene sequence of the target protein and the required unnatural amino acid information, and our team of experts will be responsible for all subsequent experimental steps, including plasmid construction, strain selection, expression condition optimization, protein purification and quality verification.

We provide the following services:

Customized unnatural amino acid incorporation: 150+ unnatural amino acids are available to meet different research needs

Protein expression and purification: Preparation of high-purity proteins from milligrams to grams

Orthogonal system construction: Design specific aaRS/tRNA orthogonal pairs for customers

Site-selective modification: Introduce functional groups at specific sites of polypeptides/proteins

Analytical verification services: Confirm incorporation efficiency and protein integrity through mass spectrometry, Western blot and other technologies

Figure 1. Service Process.

Our Advantages

• Professional team: Experienced protein engineers provide full technical support to optimize expression and purification schemes.
• High incorporation efficiency: Ensure product purity and correct modification through multiple analyses such as SDS-PAGE, mass spectrometry, and HPLC.
• Diverse selection: Supports 150+ non-natural amino acids, covering functional groups such as photocrosslinking, fluorescence, and biotin.
• One-stop service: One-stop service from gene design to protein purification

FAQs

Please contact us to provide more detailed information for evaluation.