Beyond contacts: The important role of the support region in protein complex assembly.

Protein-protein interactions (PPIs) are fundamental to nearly all cellular processes; however, elucidating the principles governing protein association into complexes remains a significant challenge. Previous studies have shown that protein-protein interfaces can be partitioned into distinct regions-core, rim, and support-that differ in hydration and residue composition. Here, we present a detailed analysis of interactions occurring within each of these interface regions. Extending beyond simple residue proximity or atomic contacts, our analysis, facilitated by our open-source software MICLOT, distinguishes among 18 different types of non-bonded interactions and evaluates their dependence on the local region. Our results demonstrate that, despite its relatively low solvent accessibility prior to complex formation, the support region contains a significant number of interactions stabilizing the interface. In particular, we find that the support promotes specific residue pair interactions, including hydrogen bonds, aromatic-aromatic and arginine stacking interactions, as well as van der Waals contacts. Furthermore, we observe that the size of the support region positively correlates with overall interface stability, and we detect differences in residue partitioning between the support and rim regions when comparing stable and transient complexes. Additionally, we introduce innovative strategies inspired by natural language processing to analyze the diversity and co-occurrence of interacting residue pairs, enabling detailed comparison of internal local organization within stable and transient interfaces. Our analysis reveals that, although pair diversity in the core and support regions falls between the interior and rim, the core and support regions exhibit interior-like pair organization in stable interfaces. Collectively, our findings emphasize the crucial role of the local environment within an interface in shaping residue interactions and underscore the support region as a key contributor to protein complex stability.