Abstract
Multivalent proteins or protein dendrimers are useful for clinical and biotechnological applications. However, assembly of chemically defined protein dendrimers is a challenging endeavor. In the past, majority of protein dendrimers have been developed on branched lysine scaffolds and are usually limited to a valency of two to four. The naturally occurring cyclodextrin (CD) scaffold composed of 6–8 glucose units offers the possibility of expanding the valency. Here we have adapted a chemoenzymatic-click strategy for displaying heptavalent peptides and large proteins on the β-cyclodextrin (β-CD) scaffold. We demonstrate that recombinant proteins (engineered with a LPXTG pentapeptide motif at the carboxy terminus), labeled with an alkyne moiety by sortase-mediated ligation, can be easily clicked on to the azide-derivatized β-cyclodextrin through the Huisgen cycloaddition reaction yielding a well-defined heptavalent display of proteins.
Highlights
Cyclodextrins (CD) are natural cyclic oligomers composed of six (α), seven (β) or eight (γ) Dglucose units linked by α- 1, 4- glycosidic bonds [1]
The successful synthesis of divalent and tetravalent proteins prompted us to explore if the SortaseClick strategy can be adapted to generate heptavalent display of large proteins on the β-CD scaffold (Fig 1)
Of chemically well-defined protein dendrimers by purely chemical methods is limited by the sequence length and associated purification complexities
Summary
Cyclodextrins (CD) are natural cyclic oligomers composed of six (α), seven (β) or eight (γ) Dglucose units linked by α- 1, 4- glycosidic bonds [1]. The noncovalent inclusion complexes of cyclodextrin have attracted a wide variety of applications for stabilization of drugs [5], solubilization of peptides and proteins, protein folding [6] etc. The hydroxyl groups on CD surface are endowed with differential reactivity and are amenable to suitable modifications with azide, alkynes, esters, sugars and other functionalities for further elaboration with a variety of macromolecules derivatized with compatible orthogonal groups [7,8,9,10,11]. CD has been found as an attractive scaffold for covalent display of peptide ligands [12,13,14].The synthesis of β-CD, symmetrically substituted with phenylalanine and cysteine residues, was reported by Ashton et al almost two decades ago [15].
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