Enthalpy arrays.
We report the fabrication of enthalpy arrays and their use to detect molecular interactions, including protein-ligand binding, enzymatic turnover, and mitochondrial respiration. Enthalpy arrays provide a universal assay methodology with no need for specific assay development such as fluorescent labeling or immobilization of reagents, which can adversely affect the interaction. Microscale technology enables the fabrication of 96-detector enthalpy arrays on large substrates. The reduction in scale results in large decreases in both the sample quantity and the measurement time compared with conventional microcalorimetry. We demonstrate the utility of the enthalpy arrays by showing measurements for two protein-ligand binding interactions (RNase A + cytidine 2'-monophosphate and streptavidin + biotin), phosphorylation of glucose by hexokinase, and respiration of mitochondria in the presence of 2,4-dinitrophenol uncoupler.
Torres
Francisco E
FE
Scripps-PARC Institute for Advanced Biomedical Sciences, Palo Alto Research Center, 3333 Coyote Hill Road, Palo Alto, CA 94304, USA.
Kuhn
Peter
P
De Bruyker
Dirk
D
Bell
Alan G
AG
Wolkin
Michal V
MV
Peeters
Eric
E
Williamson
James R
JR
Anderson
Gregory B
GB
Schmitz
Gregory P
GP
Recht
Michael I
MI
Schweizer
Sandra
S
Scott
Lincoln G
LG
Ho
Jackson H
JH
Elrod
Scott A
SA
Schultz
Peter G
PG
Lerner
Richard A
RA
Bruce
Richard H
RH
D019297:2,4-Dinitrophenol / Q000494:pharmacology; D000818:Animals; D001710:Biotin / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D019069:Cell Respiration / Q000187:drug effects; D003568:Cytidine Monophosphate / Q000737:chemistry / Q000378:metabolism*; D004867:Equipment Design; D005947:Glucose / Q000378:metabolism; D006593:Hexokinase / Q000378:metabolism; D008024:Ligands; D008929:Mitochondria, Heart / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation; D040081:Protein Array Analysis / Q000295:instrumentation / Q000379:methods*; D011485:Protein Binding; D012259:Ribonuclease, Pancreatic / Q000737:chemistry / Q000378:metabolism*; D019809:Streptavidin / Q000737:chemistry / Q000378:metabolism*; D013816:Thermodynamics
D008024:Ligands; D001710:Biotin; D019809:Streptavidin; D006593:Hexokinase; D012259:Ribonuclease, Pancreatic; D003568:Cytidine Monophosphate; D005947:Glucose; D019297:2,4-Dinitrophenol