# Carba-NAD binding activates SIR2 by reshaping conformational plasticity and rewiring long-range allosteric networks

## TL;DR

This study reveals how NAD⁺ activates SIR2 proteins by altering their structure and communication networks, offering new drug targets for aging-related diseases.

## Contribution

The study identifies a 'core-locking with peripheral-release' dynamic mode and relay-type residues that rewire allosteric communication in SIR2 upon NAD⁺ binding.

## Key findings

- Cofactor binding rigidifies the β1–α2 loop while increasing flexibility in distal regions.
- Relay residues like Pro214 and Thr224 rewire the allosteric network into a multi-node relay-style system.
- A druggable cavity overlapping with relay residues is identified as a potential target for longevity-promoting activators.

## Abstract

Allosteric regulation enables proteins to couple local structural changes to distal functional outcomes, yet the underlying mechanisms often remain difficult to fully decipher. Using yeast SIR2, an NAD ⁺ -dependent deacetylase, as a model system, this study systematically elucidates how cofactor binding reshapes its conformational dynamics and internal communication network. Through multiple 3-μs molecular dynamics simulations combined with a graph-based deep learning model (Neural Relational Inference), we identify a highly reproducible characteristic response across independent replicates: the β1–α2 loop near the active site undergoes pronounced rigidification, whereas several distal structural modules exhibit concomitant increases in flexibility, together forming a “core-locking with peripheral-release” dynamic mode. Further signal-pathway analysis reveals that the local and distal conformational changes are not independent; instead, they are interconnected through newly identified “relay-type” residues such as Pro214 and Thr224. These residues act as bridges, converting the previously β1–α2-centered centralized network into a relay-style network coordinated by multiple nodes, thereby establishing a continuous and directionally coherent allosteric cascade. Beyond mechanistic insights, we also identify a distal cavity spatially overlapping with key relay residues, whose physicochemical properties meet the criteria of druggable pockets. This structural convergence suggests that future small-molecule allosteric activators may exploit this intrinsic communication pathway to mimic or amplify the regulatory effects of the cofactor NAD ⁺. Given that NAD⁺ levels decline with aging, this cavity provides a rational target for designing longevity-promoting allosteric activators.

Sirtuins (SIR2) are a family of longevity-associated proteins that play key roles in aging, metabolism, and genome stability, with their enzymatic activity dependent on the cofactor NAD ⁺. Although their static structures have been resolved, how NAD⁺ triggers activation in distal regions remains unclear. In this study, we combine long-timescale molecular dynamics simulations with Neural Relational Inference (NRI) to elucidate the intramolecular signal transduction mechanism following the binding of the NAD⁺ analog Carba-NAD. We find that cofactor binding stabilizes the catalytic region while increasing the flexibility of distal regions, thereby reshaping the energy landscape and promoting the formation of activation-prone conformations. More importantly, the network of residue–residue interactions is reorganized, with key residues acting as bridges that facilitate long-range signal propagation. These findings provide new insights into the molecular regulatory mechanism of SIR2 and offer potential strategies for drug development targeting aging-related diseases.