Protein adaptors assemble functional proteins on DNA scaffolds

Abstract

The methods applied for assembling proteins of interest on DNA scaffolds were categorized and the recent advance of DNA-binding adaptor mediated assembly of proteins on the DNA scaffolds is discussed.

Molecular underpinnings of ssDNA specificity by Rep HUH-endonucleases and implications for HUH-tag multiplexing and engineering

“…‘HUH-tag’ fusion proteins are emerging as a versatile bioconjugation platform to covalently link proteins to DNA, combining the diverse functionality of proteins with the programmability of DNA ( 4 ). HUH-tag applications have permeated into technologies such as DNA origami scaffolded protein assembly ( 5–8 ), receptor-specific cell targeting by adeno-associated virus ( 9 ), aptamer-based sandwich detection ( 10 ), directed nanoparticle drug-delivery via DNA aptamers ( 11 ), and CRISPR–Cas9 genome engineering ( 12 , 13 ), mainly due to their ability to form robust covalent adducts under physiologic conditions. Rather than relying on expensive nucleic acid modifications such as the SNAP-tag ( 14 ), CLIP-tag ( 15 ) and HALO-tag ( 16 ) systems, HUH-tags rely on an inherent ssDNA binding moiety that promotes the catalysis of a transesterification reaction resulting in a stable phosphotyrosine adduct ( 1 ).…”

Site-selective and inducible acylation of thrombin using aptamer-catalyst conjugates

“…11,12 Theoretically, this principle can also be applied using protein-binding aptamers, [13][14][15][16] but this has not yet been shown. This is unfortunate as aptamers -which are nucleotidebased receptor molecules that bind a range of specific targets from small molecules to proteins [17][18][19] -are not only conveniently synthesized 20 and functionalized, 21 but sitespecifically bind to a protein of interest, 18 can easily be included in more complex systems [22][23][24] that, for example, facilitate ON/OFF switchable characteristics. 25,26 As such, aptamers offer a potentially rich platform for the site-selective modification of native proteins (Fig.…”

Enhanced enzymatic activity exerted by a packed assembly of a single type of enzyme

“…[15][16][17][18] In particular, DNA origami-based scaffolds enable the assembly of several different enzymes to address the effect of inter-enzyme distances on sequential reactions. [19][20][21][22][23][24] Here we apply a protein adaptor 25 to assemble a single type of monomeric enzyme, carbonic anhydrase, on a DNA scaffold in a packed state with less than a few nanometers inter-enzyme distance or dispersed state and show that the esterase reaction proceeds faster in the packed than in the dispersed state. In the packed assembly, the reaction is accelerated more prominently for substrates with higher hydrophobicity and is more tolerant of inhibitors.…”