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TitleMechanism of auto-inhibition and activation of Mec1 checkpoint kinase.
Journal, issue, pagesNat Struct Mol Biol, Vol. 28, Issue 1, Page 50-61, Year 2021
Publish dateNov 9, 2020
AuthorsElias A Tannous / Luke A Yates / Xiaodong Zhang / Peter M Burgers /
PubMed AbstractIn response to DNA damage or replication fork stalling, the basal activity of Mec1 is stimulated in a cell-cycle-dependent manner, leading to cell-cycle arrest and the promotion of DNA repair. Mec1 ...In response to DNA damage or replication fork stalling, the basal activity of Mec1 is stimulated in a cell-cycle-dependent manner, leading to cell-cycle arrest and the promotion of DNA repair. Mec1 dysfunction leads to cell death in yeast and causes chromosome instability and embryonic lethality in mammals. Thus, ATR is a major target for cancer therapies in homologous recombination-deficient cancers. Here we identify a single mutation in Mec1, conserved in ATR, that results in constitutive activity. Using cryo-electron microscopy, we determine the structures of this constitutively active form (Mec1(F2244L)-Ddc2) at 2.8 Å and the wild type at 3.8 Å, both in complex with Mg-AMP-PNP. These structures yield a near-complete atomic model for Mec1-Ddc2 and uncover the molecular basis for low basal activity and the conformational changes required for activation. Combined with biochemical and genetic data, we discover key regulatory regions and propose a Mec1 activation mechanism.
External linksNat Struct Mol Biol / PubMed:33169019 / PubMed Central
MethodsEM (single particle)
Resolution2.82 - 4.3 Å
Structure data

11050

6z2w

EMDB-11050, PDB-6z2w:
Mec1-Ddc2 (F2244L mutant) in complex with Mg AMP-PNP
Method: EM (single particle) / Resolution: 2.82 Å

11051

6z2x

EMDB-11051, PDB-6z2x:
Mec1-Ddc2 (F2244L mutant) in complex with Mg AMP-PNP (State II)
Method: EM (single particle) / Resolution: 3.2 Å

11055

6z3a

EMDB-11055, PDB-6z3a:
Mec1-Ddc2 (wild-type) in complex with AMP-PNP
Method: EM (single particle) / Resolution: 3.8 Å

EMDB-11056:
Mec1-Ddc2 (wild-type) apo
Method: EM (single particle) / Resolution: 4.3 Å

Chemicals

ChemComp-ANP:
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / AMP-PNP, energy-carrying molecule analogue*YM

ChemComp-ZN:
Unknown entry

ChemComp-MG:
Unknown entry

ChemComp-HOH:
WATER

Source
  • saccharomyces cerevisiae s288c (yeast)
  • Saccharomyces cerevisiae (brewer's yeast)
KeywordsHYDROLASE / serine/threonine protein kinase / complex / DNA damage response / checkpoint control

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