کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1358496 | 981345 | 2012 | 8 صفحه PDF | دانلود رایگان |
Covalent protein–oligodeoxynucleotide (protein–ODN) conjugates are useful in a number of biological applications, but synthesizing discrete conjugates—where the connection between the two components is at a defined location in both the protein and the ODN—under mild conditions with significant yield can be a challenge. In this article, we demonstrate a strategy for synthesizing discrete protein–ODN conjugates using strain-promoted azide–alkyne [3+2] cycloaddition (SPAAC, a copper-free ‘click’ reaction). Azide-functionalized proteins, prepared by enzymatic prenylation of C-terminal CVIA tags with synthetic azidoprenyl diphosphates, were ‘clicked’ to ODNs that had been modified with a strained dibenzocyclooctyne (DIBO–ODN). The resulting protein–ODN conjugates were purified and characterized by size-exclusion chromatography and gel electrophoresis. We find that the yields and reaction times of the SPAAC bioconjugation reactions are comparable to those previously reported for copper-catalyzed azide–alkyne [3+2] cycloaddition (CuAAC) bioconjugation, but require no catalyst. The same SPAAC chemistry was used to immobilize azide-modified proteins onto surfaces, using surface-bound DIBO–ODN as a heterobifunctional linker. Cu-free click bioconjugation of proteins to ODNs is a simple and versatile alternative to Cu-catalyzed click methods.
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Journal: Bioorganic & Medicinal Chemistry - Volume 20, Issue 14, 15 July 2012, Pages 4532–4539