Copper-Free Azide-Alkyne Cycloaddition for Peptide Modification of Alginate Hydrogels

Era Jain, Sydney Neal, Hannah Graf, Xiaohong Tan, Rama Balasubramaniam, Nathaniel Huebsch

Research output: Contribution to journalArticlepeer-review

14 Scopus citations


Alginate, a biocompatible polymer naturally derived from algae, is widely used as a synthetic analogue of the extracellular matrix in tissue engineering. Integrin-binding peptide motifs, including RGD, a derivative of fibronectin, are typically grafted to the alginate polymer through carbodiimide reactions between peptide amines and alginate uronic acids. However, lack of chemo-selectivity of carbodiimide reactions can lead to side reactions that lower peptide bioactivity. To overcome these limitations, we developed an approach for copper-free, strain-promoted azide-alkyne cycloaddition (SPAAC)-mediated conjugation of azide-modified adhesive peptides (azido-cyclo-RGD, Az-cRGD) onto alginate. Successful conjugation of azide-reactive cyclooctynes onto alginates using a heterobifunctional crosslinker was confirmed by azido-coumarin fluorescent assay, NMR, and through click reactions with azide-modified fluorescent probes. Compared to cyclo-RGD peptides directly conjugated to alginate polymers with standard carbodiimide chemistry, Az-cyclo-RGD peptides exhibited higher bioactivity, as demonstrated by cell adhesion and proliferation assays. Finally, Az-cRGD peptides enhanced the effects of recombinant bone morphogenetic proteins on inducing osteogenesis of osteoblasts and bone marrow stromal stem cells in 3D alginate gels. SPAAC-mediated click approaches for peptide-alginate bioconjugation overcome the limitations of previous alginate bioconjugation approaches and potentially expand the range of ligands that can be grafted to alginate polymers for tissue engineering applications.

Original languageEnglish (US)
Pages (from-to)1229-1237
Number of pages9
JournalACS Applied Bio Materials
Issue number2
StatePublished - Feb 15 2021
Externally publishedYes


  • alginate hydrogels
  • bone marrow stromal cells
  • click chemistry
  • extracellular matrix (ECM)
  • integrin

ASJC Scopus subject areas

  • General Chemistry
  • Biochemistry, medical
  • Biomedical Engineering
  • Biomaterials


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