5XZV

Crystal structure of Rad53 1-466 in complex with AMP-PNP

Summary for 5XZV

• Entry DOI: 10.2210/pdb5xzv/pdb
• Descriptor: Serine/threonine-protein kinase RAD53, PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER (2 entities in total)
• Functional Keywords: serine/threonine-protein kinase, fha domain, checkpoint kinase, transferase
• Biological source: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (Baker's yeast)
• Cellular location: Nucleus : P22216
• Total number of polymer chains: 2
• Total formula weight: 106025.33

Authors

Weng, J.H.,Tsai, M.D. (deposition date: 2017-07-14, release date: 2017-10-11, Last modification date: 2024-03-27)

Primary citation

Chen, E.S.,Weng, J.H.,Chen, Y.H.,Wang, S.C.,Liu, X.X.,Huang, W.C.,Matsui, T.,Kawano, Y.,Liao, J.H.,Lim, L.H.,Bessho, Y.,Huang, K.F.,Wu, W.J.,Tsai, M.D.
Phospho-Priming Confers Functionally Relevant Specificities for Rad53 Kinase Autophosphorylation
Biochemistry, 56:5112-5124, 2017

PubMed Abstract: The vast majority of in vitro structural and functional studies of the activation mechanism of protein kinases use the kinase domain alone. Well-demonstrated effects of regulatory domains or allosteric factors are scarce for serine/threonine kinases. Here we use a site-specifically phosphorylated SCD1-FHA1-kinase three-domain construct of the serine/threonine kinase Rad53 to show the effect of phospho-priming, an in vivo regulatory mechanism, on the autophosphorylation intermediate and specificity. Unphosphorylated Rad53 is a flexible monomer in solution but is captured in an asymmetric enzyme:substrate complex in crystal with the two FHA domains separated from each other. Phospho-priming induces formation of a stable dimer via intermolecular pT-FHA binding in solution. Importantly, autophosphorylation of unprimed and phospho-primed Rad53 produced predominantly inactive pS350-Rad53 and active pT354-Rad53, respectively. The latter mechanism was also demonstrated in vivo. Our results show that, while Rad53 can display active conformations under various conditions, simulation of in vivo regulatory conditions confers functionally relevant autophosphorylation.

PubMed: 28858528
DOI: 10.1021/acs.biochem.7b00689

Experimental method

X-RAY DIFFRACTION (3.1 Å)