Abstract
Proteins with tandem repeats have essential physical or biological roles in cells and have been widely investigated as biomaterials or vaccines. Chemically derived proteins with tandem repeats would be beneficial for research, owing to their accurate structures, possibly with precise modifications, produced by chemical synthesis. Traditional protein synthesis often leads to severe handling loss due to multiple ligations and HPLC purifications. To improve the protein synthesis efficiency, we developed a purification/protection handle consisting of a His6 tag and a photo-labile linker. This handle has great potential to facilitate purification with immobilized metal affinity chromatography techniques and also provides orthogonal protection for N-terminal Cys. The synthesis of the model proteins Muc1 and antifreeze glycoprotein mimics shows that the handle decreases the requirement for HPLC and enables both convergent and sequential assembly of peptide segments.
Highlights
Proteins with tandem repeats consisting of linear arrays of identical or similar repeating sequences have been shown to have important physical or biological functions.[1,2,3] According to a genome analysis, about 30% of human proteins contain tandem repeats.[4]
With or without further desulfurization, ligations can be achieved at sites with cysteine, alanine, or other residues if a beta-thio amino acid is used in the ligation step.[51,52,53,54]
We developed a useful method using a removable puri cation/ protection handle to achieve fast and efficient synthesis of proteins with tandem repeats. This method may facilitate the synthesis of more complex proteins with tandem repeats
Summary
Proteins with tandem repeats consisting of linear arrays of identical or similar repeating sequences have been shown to have important physical or biological functions.[1,2,3] According to a genome analysis, about 30% of human proteins contain tandem repeats.[4]. This handle has great potential to facilitate purification with immobilized metal affinity chromatography techniques and provides orthogonal protection for N-terminal Cys. The synthesis of the model proteins Muc[1] and antifreeze glycoprotein mimics shows that the handle decreases the requirement for HPLC and enables both convergent and sequential assembly of peptide segments.
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