Haq Tamanna, Richards Mark W, Burgess Selena G, Gallego Pablo, Yeoh Sharon, O'Regan Laura, Reverter David, Roig Joan, Fry Andrew M, Bayliss Richard
Department of Biochemistry, University of Leicester, Lancaster Road, Leicester LE1 9HN, UK.
Cancer Research UK Leicester Centre, Leicester LE1 9HN, UK.
Nat Commun. 2015 Nov 2;6:8771. doi: 10.1038/ncomms9771.
Mitotic spindle assembly requires the regulated activities of protein kinases such as Nek7 and Nek9. Nek7 is autoinhibited by the protrusion of Tyr97 into the active site and activated by the Nek9 non-catalytic C-terminal domain (CTD). CTD binding apparently releases autoinhibition because mutation of Tyr97 to phenylalanine increases Nek7 activity independently of Nek9. Here we find that self-association of the Nek9-CTD is needed for Nek7 activation. We map the minimal Nek7 binding region of Nek9 to residues 810-828. A crystal structure of Nek7(Y97F) bound to Nek9(810-828) reveals a binding site on the C-lobe of the Nek7 kinase domain. Nek7(Y97F) crystallizes as a back-to-back dimer between kinase domain N-lobes, in which the specific contacts within the interface are coupled to the conformation of residue 97. Hence, we propose that the Nek9-CTD activates Nek7 through promoting back-to-back dimerization that releases the autoinhibitory tyrosine residue, a mechanism conserved in unrelated kinase families.
有丝分裂纺锤体组装需要蛋白激酶（如Nek7和Nek9）的调控活性。Nek7因酪氨酸97突入活性位点而处于自身抑制状态，并被Nek9非催化性C末端结构域（CTD）激活。CTD结合显然会解除自身抑制，因为将酪氨酸97突变为苯丙氨酸会独立于Nek9增加Nek7的活性。在这里，我们发现Nek9-CTD的自缔合是Nek7激活所必需的。我们将Nek9与Nek7结合的最小区域定位到810-828位氨基酸残基。Nek7（Y97F）与Nek9（810-828）结合的晶体结构揭示了Nek7激酶结构域C叶上的一个结合位点。Nek7（Y97F）以激酶结构域N叶之间背靠背二聚体的形式结晶，其中界面内的特定接触与97位残基的构象相关联。因此，我们提出Nek9-CTD通过促进背靠背二聚化来激活Nek7，从而释放自抑制性酪氨酸残基，这是一种在不相关激酶家族中保守的机制。
