title: Electronic measurements of single-molecule catalysis by cAMP-dependent protein kinase A.
abstract: Single-molecule studies of enzymes open a window into their dynamics and kinetics. A single molecule of the catalytic domain of cAMP-dependent protein kinase A (PKA) was attached to a single-walled carbon nanotube device for long-duration monitoring. The electronic recording clearly resolves substrate binding, ATP binding, and cooperative formation of PKA's catalytically functional, ternary complex. Using recordings of a single PKA molecule extending over 10 min and tens of thousands of binding events, we determine the full transition probability matrix and conversion rates governing formation of the apo, intermediate, and closed enzyme configurations. We also observe kinetic rates varying over 2 orders of magnitude from one second to another. Anti-correlation of the on and off rates for PKA binding to the peptide substrate, but not ATP, demonstrates that regulation of enzyme activity results from altering the stability of the PKA-substrate complex, not its binding to ATP. The results depict a highly dynamic enzyme offering dramatic possibilities for regulated activity, an attribute useful for an enzyme with crucial roles in cell signaling.
authors: Department of Physics and Astronomy, University of California, Irvine, California 92697, USA.
authors: Patrick C
authors: PC
authors: Sims
authors: Issa S
authors: IS
authors: Moody
authors: Yongki
authors: Y
authors: Choi
authors: Chengjun
authors: C
authors: Dong
authors: Mariam
authors: M
authors: Iftikhar
authors: Brad L
authors: BL
authors: Corso
authors: O Tolga
authors: OT
authors: Gul
authors: Philip G
authors: PG
authors: Collins
authors: Gregory A
authors: GA
authors: Weiss
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D002384:Catalysis; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D007700:Kinetics; D037742:Nanotubes, Carbon
chemical_list: D037742:Nanotubes, Carbon; D000255:Adenosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases
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