Protein Kinase A Catalytic Subunit Primed for Action: Time-Lapse Crystallography of Michaelis Complex Formation

Journal Article · Tue Dec 01 00:00:00 EST 2015 · Structure

DOI:https://doi.org/10.1016/j.str.2015.10.005· OSTI ID:1334200

Das, Amit; Gerlits, Oksana; Parks, Jerry M.; Langan, Paul; Kovalevsky, Andrey; Heller, William T.

The catalytic subunit of the cyclic AMP-dependent protein kinase A (PKAc) catalyzes the transfer of the γ-phosphate of bound Mg₂ATP to a serine or threonine residue of a protein substrate. Here, time-lapse X-ray crystallography was used to capture a series of complexes of PKAc with an oligopeptide substrate and unreacted Mg₂ATP, including the Michaelis complex, that reveal important geometric rearrangements in and near the active site preceding the phosphoryl transfer reaction. Contrary to the prevailing view, Mg²⁺ binds first to the M1 site as a complex with ATP and is followed by Mg²⁺ binding to the M2 site. Furthermore, the target serine hydroxyl of the peptide substrate rotates away from the active site toward the bulk solvent, which breaks the hydrogen bond with D166. In conclusion, the serine hydroxyl of the substrate rotates back toward D166 to form the Michaelis complex with the active site primed for phosphoryl transfer.

• Research Organization: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)
• Sponsoring Organization: USDOE Office of Science (SC), Basic Energy Sciences (BES)
• Grant/Contract Number: AC05-00OR22725
• OSTI ID: 1334200
• Alternate ID(s): OSTI ID: 1235817; OSTI ID: 1250913
• Journal Information: Structure, Journal Name: Structure Vol. 23 Journal Issue: 12; ISSN 0969-2126
• Publisher: Elsevier
• Country of Publication: United Kingdom
• Language: English

Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY