DNA sensing by electrocatalysis with hemoglobin.
Electrocatalysis offers a means of electrochemical signal amplification, yet in DNA-based sensors, electrocatalysis has required high-density DNA films and strict assembly and passivation conditions. Here, we describe the use of hemoglobin as a robust and effective electron sink for electrocatalysis in DNA sensing on low-density DNA films. Protein shielding of the heme redox center minimizes direct reduction at the electrode surface and permits assays on low-density DNA films. Electrocatalysis with methylene blue that is covalently tethered to the DNA by a flexible alkyl chain linkage allows for efficient interactions with both the base stack and hemoglobin. Consistent suppression of the redox signal upon incorporation of a single cytosine-adenine (CA) mismatch in the DNA oligomer demonstrates that both the unamplified and the electrocatalytically amplified redox signals are generated through DNA-mediated charge transport. Electrocatalysis with hemoglobin is robust: It is stable to pH and temperature variations. The utility and applicability of electrocatalysis with hemoglobin is demonstrated through restriction enzyme detection, and an enhancement in sensitivity permits femtomole DNA sampling.
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
Catrina G
CG
Pheeney
Luis F
LF
Guerra
Jacqueline K
JK
Barton
D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000737:chemistry*; D004262:DNA Restriction Enzymes / Q000737:chemistry; D055664:Electrochemical Techniques / Q000379:methods*; D004566:Electrodes; D006454:Hemoglobins / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008751:Methylene Blue; D015394:Molecular Structure; D009841:Oligonucleotides / Q000235:genetics; D010084:Oxidation-Reduction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013696:Temperature
D006454:Hemoglobins; D009841:Oligonucleotides; D004247:DNA; D004262:DNA Restriction Enzymes; D008751:Methylene Blue