Glen Report 35-25: Product Review DBCO

In recent years, one type of Click Chemistry called  Strain-Promoted Azide-Alkyne Cycloaddition (SPAAC)1 has become particularly popular. The conjugation reaction is relatively fast, selective, and compatible with biological environments. The reaction involves an azide and a strained alkyne to form a stable 1,2,3-trieat to using them is that iodine oxidizer and DBCO are not compatible, which necessitates the use of our mild oxidizing agent, CSO, instead. For a 5′ addition, only one cycle of CSO oxidation is required whereas for a 3′ or internal insertion, CSO oxidation will be required throughout the whole synthesis. For those who would like to avoid the use of CSO, we also offer an NHS ester (Figure 2) that will label any amino modifier group post oligonucleotide deprotection. For example, one can  pair the NHS ester with 5′-Amino-Modifier TEG CE- Phosphoramidite or Amino-Modifier C6 dT (Figure 2) azole linkage (Figure 1), and the strained alkyne is the key chemical group that makes it all possible. 

Figure 1

Figure 1. SPAAC with the strained alkyne DBCO

Alkynes are ideally linear, and when they are incorporated into cyclic analogs, particularly smaller ones, the alkyne will have angle strain. While linear alkynes require toxic copper catalysts to facilitate conjugation, strained alkynes will spontaneously react. The formation of the triazole ring converts the higher energy, strained alkyne to an alkene, thereby relieving the ring strain.

We offer dibenzocyclooctyne, DBCO, as a modification for facilitating SPAAC with oligonucleotides. There are three phosphoramidites: 5′-DBCO-TEG Phosphoramidite, DBCO-dT-CE Phosphoramidite, and DBCO-Serinol Phosphoramidite for 5′, internal, and general oligonucleotide labeling, respectively (Figure 2).  These products work great. The only cav to give the same exact structures obtained from two of the three DBCO phosphoramidites.

Figure 2

Figure 2. DBCO products and possible amino-modifier precursors

Our DBCO products have been successfully used in a wide range of investigations. In one project, DBCO facilitated the labeling of catalase enzymes with oligonucleotides for protein-protein and protein-gold nanoparticle superlattice formation.2 Another group of researchers used DBCO to attach an RNA adaptor sequence to an mRNA 5′-cap to enable nanopore sequencing of full-length mRNA sequences.3 In a third study, DBCO allowed antisense oligonucleotides to be attached to monoclonal antibodies for leukemia-targeting.4 The resulting antibody-drug conjugate was effective in both in vitro and in vivo mouse model experiments. Finally, DBCO facilitated the engraftment of proteins onto modular DNA scaffolds for the analysis of protein-protein interactions at single-molecule resolution.5

For those considering conjugations and bioconjugations, SPAAC and DBCO should be one approach worthy of strong consideration.

Product Information

Conjugation using Click Chemistry