Hameed, U. F. S. | Search bioRxiv

bioRxiv2024

Regulation of Arabidopsis polyamine acetylation by NATA1 and NATA2

Polyamines have vital functions in organisms, including bacteria, plants, and animals, with key roles in growth, development, and stress responses. Spermine/spermidine N1-acetyl transferases (SSATs) regulate polyamine abundance by catalysing their N-acetylation, thereby reducing the pool of polyamines and producing other bioactive components. The regulatory mechanisms controlling SSAT enzymes are incompletely understood. Here, we investigate the biological role and regulation of the two SSAT isoforms present in Arabidopsis thaliana, N-ACETYLTRANSFERASE ACTIVITY (NATA) 1 and 2. We show that NATA2 is a heat-stable isoform, induced in response to heat. Intriguingly, a nata2 knockout mutation proved beneficial for growth and pathogen defence under heat stress in Arabidopsis, aligning with the stress-mitigating effect of polyamines. In contrast, the double knockout of nata1 and nata2 was lethal, highlighting the essential role of basal SSAT activity. Our numerous crystal structures of both NATAs, supported by functional assays, revealed that stress-produced acidic metabolites can selectively inhibit polyamine acetylation by occupying the NATA substrate-binding pocket. This environment-responsive regulation mechanism may allow Arabidopsis to adjust the deleterious action of NATAs under stress conditions, without eliminating the enzyme. More generally, metabolite-ensemble inhibition may be a novel paradigm for non-genetic feedback regulation of plant enzymes.

plant biology↗

bioRxiv2020

Synergy and allostery in ligand binding by HIV-1 Nef

The Nef protein of human and simian immunodeficiency viruses (HIV and SIV, respectively) boosts viral pathogenicity through its interactions with host cell proteins. Nef has a folded core domain and large flexible regions, each carrying several protein interaction sites. By combining the polyvalency intrinsic to unstructured regions with the binding selectivity and strength of a 3D folded domain, Nef can bind to many different host cell proteins, perturbing their cellular functions. For example, the combination of a linear proline-rich motif and a hydrophobic core domain surface allows Nef to increase affinity and selectivity for particular Src family SH3 domains. Here we investigated whether the interplay between Nefs flexible regions and its core domain can allosterically influence ligand selection. We found that the flexible regions can bind back to the core domain in different ways, producing distinct conformational states that alter the SH3 domain selectivity and availability of Nefs functional motifs. The resulting cross-talk might help synergising certain subsets of ligands while excluding others, promoting functionally coherent Nef-bound protein ensembles. Further, we combined proteomic and bioinformatic analyses to identify human proteins that select SH3 domains in the same way as does Nef. We found that only 2-3% of clones from a whole human fetal library displayed a Nef-like SH3 selectivity. However, in most cases this selectivity appears to be achieved by a canonical linear interaction rather than a Nef-like tertiary interaction. This analysis suggests that Nefs SH3 recognition surface has no (or marginally few) cellular counterparts, validating the Nef tertiary binding surface as a promising unique drug target.

biophysics↗