A Rationally Designed Connector for Assembly of Protein-Functionalized DNA Nanostructures

We report on the rational engineering of the binding interface of the self‐ligating HaloTag protein to generate an optimized linker for DNA nanostructures. Five amino acids positioned around the active‐site entry channel for the chlorohexyl ligand (CH) of the HaloTag protein were exchanged for posit...

Full description

Saved in:
Bibliographic Details
Published in:Chembiochem : a European journal of chemical biology 2016-06, Vol.17 (12), p.1102-1106
Main Authors: Koßmann, Katja J., Ziegler, Cornelia, Angelin, Alessandro, Meyer, Rebecca, Skoupi, Marc, Rabe, Kersten S., Niemeyer, Christof M.
Format: Article
Language:English
Subjects:
Amino acids
Binding Sites
bioconjugates
Catalytic Domain
Cytochrome P-450 Enzyme System - chemistry
Cytochrome P-450 Enzyme System - metabolism
Deoxyribonucleic acid
DNA
DNA - chemistry
DNA nanostructures
enzymes
Microscopy, Atomic Force
Nanostructures - chemistry
Oligonucleotides - chemistry
Proteins
Recombinant Proteins - chemistry
Recombinant Proteins - metabolism
self-assembly
Static Electricity
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:We report on the rational engineering of the binding interface of the self‐ligating HaloTag protein to generate an optimized linker for DNA nanostructures. Five amino acids positioned around the active‐site entry channel for the chlorohexyl ligand (CH) of the HaloTag protein were exchanged for positively charged lysine amino acids to produce the HOB (halo‐based oligonucleotide binder) protein. HOB was genetically fused with the enzyme cytochrome P450 BM3, as well as with BMR, the separated reductase domain of BM3. The resulting HOB‐fusion proteins revealed significantly improved rates in ligation with CH‐modified oligonucleotides and DNA origami nanostructures. These results suggest that the efficient self‐assembly of protein‐decorated DNA structures can be greatly improved by fine‐tuning of the electrostatic interactions between proteins and the negatively charged nucleic acid nanostructures. HOBbing proteins with DNA: Halo‐based oligonucleotide binder (HOB, right) is an oligonucleotide‐binder protein in which positively charged amino acids (blue) were incorporated around the active‐site entry channel for the chlorohexyl ligand (green) of the self‐ligating HaloTag protein (left). HOB fusion proteins bind to oligonucleotides and DNA origami with significantly improved reaction rates.
ISSN:1439-4227
1439-7633
DOI:10.1002/cbic.201600039