Packaging of the Coronavirus Ribonucleoprotein—A Symphony of Enthalpy and Entropy

ABSTRACT The coronavirus nucleocapsid protein (N) is the most abundant viral protein in infected cells and plays a central role in genome packaging, replication, and transcription. Its primary function is to organize an exceptionally large, positive‐sense RNA genome into a helical ribonucleoprotein (RNP) assembly that must be both structurally stable and dynamically accessible. In this review, we summarize our biophysical and structural studies elucidate how the severe acute respiratory syndrome coronavirus (SARS‐CoV) N protein resolves these competing demands. The N protein is shown to be modular, comprising two folded domains embedded within extensive intrinsically disordered regions, resulting in a multivalent, electrostatically driven RNA‐binding architecture. Quantitative binding analyses reveal moderate‐affinity interactions distributed across multiple sites and coupled through positive cooperativity. High‐resolution structures of the C‐terminal dimerization domain obtained by X‐ray crystallography and solution NMR reveal a oligomerization scaffold and delineate an extended helical RNA‐binding surfaces. These findings highlight a simple physical principle, namely that CoV RNP packaging is a symphony of enthalpic contribution from electrostatic interaction and entropic contribution inherent to the intrinsic disordered regions. While the enthalpic energy stabilizes the RNP structure, the entropic effect facilitates the N‐RNA interaction and structural rearrangement. Together, these studies establish a physical framework in which structural order and intrinsic disorder cooperate to enable efficient and reversible ribonucleoprotein assembly.