Nanobodies identity an activated state of the TRIB2 pseudokinase

Jamieson, S. A., Pudjihartono, M., Horne, C. R., Viloria, J. S., Dunlop, J. L., McMillan, H. D., Day, R. C., Keeshan, K. , Murphy, J. M. and Mace, P. D. (2022) Nanobodies identity an activated state of the TRIB2 pseudokinase. Structure, 30(11), 1518-1529.e5. (doi: 10.1016/j.str.2022.08.006) (PMID:36108635)

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Tribbles proteins (TRIB1–3) are pseudokinases that recruit substrates to the COP1 ubiquitin ligase. TRIB2 was the first Tribbles ortholog to be implicated as a myeloid leukemia oncogene, because it recruits the C/EBPα transcription factor for ubiquitination by COP1. Here we report identification of nanobodies that bind the TRIB2 pseudokinase domain with low nanomolar affinity. A crystal structure of the TRIB2-Nb4.103 complex identified the nanobody to bind the N-terminal lobe of TRIB2, enabling specific recognition of TRIB2 in an activated conformation that is similar to the C/EBPα-bound state of TRIB1. Characterization in solution revealed that Nb4.103 can stabilize a TRIB2 pseudokinase domain dimer in a face-to-face manner. Conversely, a distinct nanobody (Nb4.101) binds through a similar epitope but does not readily promote dimerization. In combination, this study identifies features of TRIB2 that could be exploited for the development of inhibitors and nanobody tools for future investigation of TRIB2 function.

Item Type:Articles
Additional Information:This work was supported by the Marsden Fund Council from New Zealand Government funding, managed by Royal Society Te Apārangi. J.L.D. and H.D.M. received support from University of Otago Doctoral Scholarships. J.M.M. gratefully acknowledges the support of the National Health and Medical Research Council of Australia (fellowship, 1172929; IRIISS, 9000719) and the Victorian Government Operational Infrastructure Support scheme. K.K. acknowledges the support of Blood Cancer UK (19012).
Glasgow Author(s) Enlighten ID:Keeshan, Dr Karen
Authors: Jamieson, S. A., Pudjihartono, M., Horne, C. R., Viloria, J. S., Dunlop, J. L., McMillan, H. D., Day, R. C., Keeshan, K., Murphy, J. M., and Mace, P. D.
College/School:College of Medical Veterinary and Life Sciences > School of Cancer Sciences
Journal Name:Structure
ISSN (Online):1878-4186
Published Online:14 September 2022
Copyright Holders:Crown Copyright © 2022
First Published:First published in Structure 30(11):1518-1529.e5
Publisher Policy:Reproduced in accordance with the publisher copyright policy

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