Structural and mechanistic insights into Mps1 kinase activation.
Mps1 is one of the several essential kinases whose activation is required for robust mitotic spindle checkpoint signalling. The activity of Mps1 is tightly regulated and increases dramatically during mitosis or in response to spindle damage. To understand the molecular mechanism underlying Mps1 regulation, we determined the crystal structure of the kinase domain of Mps1. The 2.7-A-resolution crystal structure shows that the Mps1 kinase domain adopts a unique inactive conformation. Intramolecular interactions between the key Glu residue in the C helix of the N-terminal lobe and the backbone amides in the catalytic loop lock the kinase in the inactive conformation. Autophosphorylation appears to be a priming event for kinase activation. We identified Mps1 autophosphorylation sites in the activation and the P+1 loops. Whereas activation loop autophosphorylation enhances kinase activity, autophosphorylation at the P+1 loop (T686) is associated with the active kinase. Mutation of T686 autophosphorylation site impairs both autophosphorylation and transphosphorylation. Furthermore, we demonstrated that phosphorylation of T676 may be a priming event for phosphorylation at T686. Finally, we identified two critical lysine residues in the loop between helices EF and F that are essential for substrate recruitment and maintaining high levels of kinase activity. Our studies reveal critical biochemical mechanisms for Mps1 kinase regulation.
Wang
Wei
W
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
Yang
Yuting
Y
Department of Biological Chemistry, University of Michigan, Ann Arbor, MI, USA.
Gao
Yuefeng
Y
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
Xu
Quanbin
Q
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
Wang
Feng
F
Department of Biological Chemistry, University of Michigan, Ann Arbor, MI, USA.
Zhu
Songcheng
S
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
Old
William
W
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
Resing
Katheryn
K
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
Ahn
Natalie
N
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.|Howard Hughes Medical Institute, University of Colorado, Boulder, CO, USA.
Lei
Ming
M
Department of Biological Chemistry, University of Michigan, Ann Arbor, MI, USA.
Liu
Xuedong
X
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO, USA.
D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases; D013379:Substrate Specificity
D018797:Cell Cycle Proteins; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; C075840:TTK protein, human