Rapid synthesis of DNA-cysteine conjugates for expressed protein ligation

@article{Niemeyer2005, abstract = {We report a rapid method for the covalent modification of commercially available amino-modified DNA oligonucleotides with a cysteine moiety. The resulting DNA-cysteine conjugates are versatile reagents for the efficient preparation of covalent DNA-protein conjugates by means of expressed protein ligation (EPL). The EPL method allows for the site-specific coupling of cysteine-modified DNA oligomers with recombinant intein-fusion proteins, the latter of which contain a C-terminal thioester enabling the mild and highly specific reaction with N-terminal cysteine compounds. We prepared a cysteine-modifier reagent in a single-step reaction which allows for the rapid and near quantitative synthesis of cysteine-DNA conjugates. The latter were ligated with the green fluorescent protein mutant EYFP, recombinantly expressed as an intein-fusion protein, allowing for the mild and selective formation of EYFP-DNA conjugates in high yields of about 60\%. We anticipate many applications of our approach, ranging from protein microarrays to the arising field of nanobiotechnology.}, author = {Lovrinovic, Marina and Niemeyer, Christof M. }, citeulike-article-id = {2401667}, doi = {10.1016/j.bbrc.2005.08.001}, journal = {Biochemical and Biophysical Research Communications}, keywords = {addons, addons-protein, origami}, month = {September}, number = {3}, pages = {943--948}, posted-at = {2008-02-20 06:53:03}, priority = {2}, title = {Rapid synthesis of DNA-cysteine conjugates for expressed protein ligation}, url = {http://dx.doi.org/10.1016/j.bbrc.2005.08.001}, volume = {335}, year = {2005} }

TY - JOUR ID - Niemeyer2005 L3 - citeulike-article-id:2401667 TI - Rapid synthesis of DNA-cysteine conjugates for expressed protein ligation JF - Biochemical and Biophysical Research Communications VL - 335 IS - 3 SP - 943 EP - 948 N2 - We report a rapid method for the covalent modification of commercially available amino-modified DNA oligonucleotides with a cysteine moiety. The resulting DNA-cysteine conjugates are versatile reagents for the efficient preparation of covalent DNA-protein conjugates by means of expressed protein ligation (EPL). The EPL method allows for the site-specific coupling of cysteine-modified DNA oligomers with recombinant intein-fusion proteins, the latter of which contain a C-terminal thioester enabling the mild and highly specific reaction with N-terminal cysteine compounds. We prepared a cysteine-modifier reagent in a single-step reaction which allows for the rapid and near quantitative synthesis of cysteine-DNA conjugates. The latter were ligated with the green fluorescent protein mutant EYFP, recombinantly expressed as an intein-fusion protein, allowing for the mild and selective formation of EYFP-DNA conjugates in high yields of about 60%. We anticipate many applications of our approach, ranging from protein microarrays to the arising field of nanobiotechnology. KW - addons KW - addons-protein KW - origami AU - Lovrinovic, Marina AU - Niemeyer, Christof PY - 2005/09/30/ UR - http://dx.doi.org/10.1016/j.bbrc.2005.08.001 ER -