S T R U C T U R A L B I O L O G Y

S C I E N T I F I C H I G H L I G H T S

4 2 H I G H L I G H T S 2 0 2 2 I

presence of the SSU but deleterious in its absence caused a dependence on it [3]. Rubiscos that accumulated such substitutions formed supramolecular fibrils without the SSU (Figure 32e). These substitutions cause assembly into insoluble fibrils but do not improve Rubisco s kinetic parameters in any way. Substitutions that made this interaction essential for Rubisco were therefore non- adaptive and accumulated purely because they incurred no negative costs.

These findings suggest that natural selection can fix protein-protein interactions for their beneficial effects and that such interactions can subsequently become essential via non-adaptive evolution. Furthermore, the studied

ancestral Rubiscos show that high-specificity arose early in Rubisco s history, when limited O2 was present [4]. This illuminates a key event in the history of life on Earth. It also reveals surprising things about how specificity is encoded in Rubisco, which can inform engineering ventures. Instead of being determined solely by changes in the large subunit, the recruitment of a novel interaction partner opened the way for evolution to improve Rubisco s specificity.

Small accessory subunits can be powerful modulators for the evolution of proteins. This can help access functions that are otherwise not accessible but can equally lead to non-adaptive dependencies that evolve through a shifted accessible sequence space.

PRINCIPAL PUBLICATION AND AUTHORS

Evolution of increased complexity and specificity at the dawn of form I Rubiscos, L. Schulz (a), Z. Guo (b), J. Zarzycki (a), W. Steinchen (c,d), J.M. Schuller (c,d), T. Heimerl (c,d), S. Prinz (e), O. Mueller-Cajar (b), T.J. Erb (a,c), G.K.A. Hochberg (a,c,d), Science 378, 155-160 (2022); https:/doi.org/10.1126/science.abq1416 (a) Max Planck Institute for Terrestrial Microbiology, Marburg (Germany) (b) Nanyang Technological University Singapore (Singapore) (c) Center for Synthetic Microbiology (SYNMIKRO), Marburg (Germany) (d) Philipps University Marburg (Germany) (e) Max Planck Institute for Biophysics, Frankfurt (Germany)

REFERENCES

[1] T.J. Erb et al., Curr. Opin. Biotech. 49, 100-107 (2018). [2] G.K.A Hochberg et al., Ann. Rev. Biophys. 46, 247 (2017). [3] G.K.A. Hochberg et al., Nature 588 (7838), 503-508 (2020). [4] P.M. Shih et al., Nat. Commun. 10382 (2016).

Structural basis of reactivation of oncogenic p53 mutants by a small molecule: methylene quinuclidinone (MQ) In response to cellular stress, the tumour suppressor p53 activates genes critical for cancer prevention. In more than 50% of human cancers, mutations in p53 lead to its inactivation. A small molecule, methylene quinuclidinone (MQ), reactivates mutant p53. Crystal structures of p53 mutants bound to MQ reveal the reactivation mechanism of cancer-related p53.

The p53 protein is a tumour suppressor that, in response to cellular stress, acts as a transcription factor by binding as a tetramer (a dimer of dimers) to a wide range of DNA response elements, activating various events including DNA repair, cell-cycle arrest, senescence or apoptosis, all critical to cancer prevention. The human p53 protein is 393 residues long and contains several major functional domains, of which the N-terminus contains a transactivation domain, the core domain contains a sequence-specific DNA binding domain (p53DBD) and the C-terminus incorporates tetramerisation and regulatory domains. Of these domains, only the DNA-binding and the oligomerisation domains are structured, whereas other parts of the protein are intrinsically disordered.

In more than half of human cancers, mutations are observed in the TP53 gene, leading to loss of wild-type p53 function. Over 90% of the tumorigenic mutations are found in the DNA-binding domain, spanning nearly 200 residues (94 293). Among these mutations, six are referred as hotspots due to their high frequency (nearly 30%) in various types of cancer. Mutations at positions R248 and R273, referred as DNA-contact mutations, lead to loss of direct p53-DNA interactions. Mutations at positions R175, G245, R249 and R282, referred as structural mutations, lower p53 stability and modify its folding state in a mutation-specific manner from local distortions to global denaturation.

Because of its key role in cancer development, mutant p53 has been a target for restoring wild-type function to mutant p53 by small drug molecules. Among the most clincially advanced, eprenetapopt (APR-246/PRIMA-1MET) is a small molecule that, following conversion into its biologically active compound, methylene quinuclidinone (MQ), reactivates mutant p53 by binding covalently to cysteine residues in p53. Two enantiomers may be formed upon MQ binding to a cysteine thiol group. This reaction is highly reversible; hence, the selection of either enantiomer at a specific cysteine residue varies, depending on stabilising interactions with the molecular surrounding.