TY - JOUR
T1 - Hidden Multivalency in Phosphatase Recruitment by a Disordered AKAP Scaffold
T2 - Hidden multivalency in a disordered scaffold
AU - Watson, Matthew
AU - Almeida, Teresa B.
AU - Ray, Arundhati
AU - Hanack, Christina
AU - Elston, Rory
AU - Btesh, Joan
AU - McNaughton, Peter A.
AU - Stott, Katherine
N1 - Funding Information:
We thank Peter Schuck, Marko Hyvönen and Anna Git for helpful discussions, the NMR and Biophysics Facilities in the Department of Biochemistry, University of Cambridge for access to instrumentation, and Dermot Cooper for the Calmodulin plasmid. This work was supported by the Biotechnology and Biological Sciences Research Council (BB/N022181/1 and BB/T015403/1).
Publisher Copyright:
© 2022 The Author(s)
PY - 2022/8/30
Y1 - 2022/8/30
N2 - Disordered scaffold proteins provide multivalent landing pads that, via a series of embedded Short Linear Motifs (SLiMs), bring together the components of a complex to orchestrate precise spatial and temporal regulation of cellular processes. One such protein is AKAP5 (previously AKAP79), which contains SLiMs that anchor PKA and Calcineurin, and recruit substrate (the TRPV1 receptor). Calcineurin is anchored to AKAP5 by a well-characterised PxIxIT SLiM. Here we show, using a combination of biochemical and biophysical approaches, that the Calcineurin PxIxIT-binding groove also recognises several hitherto unknown lower-affinity SLiMs in addition to the PxIxIT motif. We demonstrate that the assembly is in reality a complex system with conserved SLiMs spanning a wide affinity range. The capture is analogous to that seen for many DNA-binding proteins that have a weak non-specific affinity for DNA outside the canonical binding site, but different in that it involves (i) two proteins, and (ii) hydrophobic rather than electrostatic interactions. It is also compatible with the requirement for both stable anchoring of the enzyme and responsive downstream signalling. We conclude that the AKAP5 C-terminus is enriched in lower-affinity/mini-SLiMs that, together with the canonical SLiM, maintain a structurally disordered but tightly regulated signalosome.
AB - Disordered scaffold proteins provide multivalent landing pads that, via a series of embedded Short Linear Motifs (SLiMs), bring together the components of a complex to orchestrate precise spatial and temporal regulation of cellular processes. One such protein is AKAP5 (previously AKAP79), which contains SLiMs that anchor PKA and Calcineurin, and recruit substrate (the TRPV1 receptor). Calcineurin is anchored to AKAP5 by a well-characterised PxIxIT SLiM. Here we show, using a combination of biochemical and biophysical approaches, that the Calcineurin PxIxIT-binding groove also recognises several hitherto unknown lower-affinity SLiMs in addition to the PxIxIT motif. We demonstrate that the assembly is in reality a complex system with conserved SLiMs spanning a wide affinity range. The capture is analogous to that seen for many DNA-binding proteins that have a weak non-specific affinity for DNA outside the canonical binding site, but different in that it involves (i) two proteins, and (ii) hydrophobic rather than electrostatic interactions. It is also compatible with the requirement for both stable anchoring of the enzyme and responsive downstream signalling. We conclude that the AKAP5 C-terminus is enriched in lower-affinity/mini-SLiMs that, together with the canonical SLiM, maintain a structurally disordered but tightly regulated signalosome.
KW - A-kinase anchoring protein (AKAP)
KW - Calcineurin
KW - intrinsically disordered protein
KW - multivalency
KW - short linear motif (SLiM)
UR - http://www.scopus.com/inward/record.url?scp=85132877816&partnerID=8YFLogxK
U2 - 10.1016/j.jmb.2022.167682
DO - 10.1016/j.jmb.2022.167682
M3 - Article
AN - SCOPUS:85132877816
SN - 0022-2836
VL - 434
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 16
M1 - 167682
ER -