Orthogonal enzymatic reactions for the assembly of proteins at electrode addresses.
The ability to interface proteins to device surfaces is important for a range of applications. Here, we enlist the unique capabilities of enzymes and biologically derived polymers to assemble target proteins to electrode addresses. First, the stimuli-responsive aminopolysaccharide chitosan is directed to assemble at the electrode address in response to electrode-imposed signals. The electrodeposited chitosan film serves as the biodevice interface for subsequent protein assembly. Next, tyrosinase is used to catalyze grafting of a protein or peptide tether to the chitosan film. Finally, microbial transglutaminase (mTG) catalyzes the assembly of target proteins to the tether. mTG covalently links proteins through their glutamine (Gln) and lysine (Lys) residues. Since Gln and Lys residues of globular proteins are often inaccessible to mTG, we engineered our target proteins to have fusion tags with added Gln or Lys residues. This assembly method employs the electrical signal to confer spatial selectivity (during chitosan electrodeposition) and employs the enzymes to confer chemical selectivity (i.e., amino acid residue selectivity). Further, this method is mild, since no reactive reagents or protection steps are required, and all steps are performed in aqueous solution. These results demonstrate the potential for employing biological materials and mechanisms to biofabricate the biodevice interface.
Yang
Xiaohua
X
Center for Biosystems Research, University of Maryland Biotechnology Institute, 5115 Plant Sciences Building, College Park, Maryland 20742, USA.
Shi
Xiao-Wen
XW
Liu
Yi
Y
Bentley
William E
WE
Payne
Gregory F
GF
D001483:Base Sequence; D055162:Biocatalysis; D048271:Chitosan / Q000737:chemistry; D017931:DNA Primers; D004566:Electrodes; D005780:Gelatin / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000737:chemistry; D008856:Microscopy, Fluorescence; D014442:Monophenol Monooxygenase / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D011503:Transglutaminases / Q000737:chemistry*
D017931:DNA Primers; D011506:Proteins; D049452:Green Fluorescent Proteins; D005780:Gelatin; D048271:Chitosan; D014442:Monophenol Monooxygenase; D011503:Transglutaminases