Quantum dot peptide biosensors for monitoring caspase 3 proteolysis and calcium ions.
The nanoscale size and unique optical properties of semiconductor quantum dots (QDs) have made them attractive as central photoluminescent scaffolds for a variety of biosensing platforms. In this report we functionalize QDs with dye-labeled peptides using two different linkage chemistries to yield Förster resonance energy transfer (FRET)-based sensors capable of monitoring either enzymatic activity or ionic presence. The first sensor targets the proteolytic activity of caspase 3, a key downstream effector of apoptosis. This QD conjugate utilized carbodiimide chemistry to covalently link dye-labeled peptide substrates to the terminal carboxyl groups on the QD's surface hydrophilic ligands in a quantitative manner. Caspase 3 cleaved the peptide substrate and disrupted QD donor-dye acceptor FRET providing signal transduction of enzymatic activity and allowing derivation of relevant Michaelis-Menten kinetic descriptors. The second sensor was designed to monitor Ca2+ ions that are ubiquitous in many biological processes. For this sensor, Cu+-catalyzed [3 + 2] azide-alkyne cycloaddition was exploited to attach a recently developed azide-functionalized CalciumRuby-Cl indicator dye to a cognate alkyne group present on the terminus of a modified peptide. The labeled peptide also expressed a polyhistidine sequence, which facilitated its subsequent metal-affinity coordination to the QD surface establishing the final FRET sensing construct. Adding exogenous Ca2+ to the sensor solution increased the dyes fluorescence, altering the donor-acceptor emission ratio and manifested a dissociation constant similar to that of the native dye. These results highlight the potential for combining peptides with QDs using different chemistries to create sensors for monitoring chemical compounds and biological processes.
Prasuhn
Duane E
DE
Center for Bio/Molecular Science and Engineering, Code 6900, U.S. Naval Research Laboratory, 4555 Overlook Ave., S.W.,Washington, D.C. 20375, USA.
Feltz
Anne
A
Blanco-Canosa
Juan B
JB
Susumu
Kimihiro
K
Stewart
Michael H
MH
Mei
Bing C
BC
Yakovlev
Aleksey V
AV
Loukov
Christina
C
Mallet
Jean-Maurice
JM
Oheim
Martin
M
Dawson
Philip E
PE
Medintz
Igor L
IL
D000595:Amino Acid Sequence; D015374:Biosensing Techniques / Q000379:methods*; D002118:Calcium / Q000032:analysis*; D053148:Caspase 3 / Q000378:metabolism*; D004738:Engineering; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D045663:Quantum Dots
D005456:Fluorescent Dyes; D010455:Peptides; D053148:Caspase 3; D002118:Calcium