Structural and functional insights into I?B-?/HIV-1Tat interaction

Protein-protein interactions play fundamental roles in physiological and pathological biological processes. The characterization of the structural determinants of protein-protein recognition represents an important step for the development of molecular entities able to modulate these interactions. We have recently found that I?B-? (nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha) blocks the HIV-1 expression and replication in a NF-?B-independent manner by directly binding to the virus-encoded Tat transactivator. Here, we report the evaluation of the entity of binding of I?B-? to Tat through in vitro Surface Plasmon Resonance assay. Moreover, by designing and characterizing a set of peptides of the C-terminus region of I?B-?, we show that the peptide corresponding to the I?B-? sequence 262-287 was able to bind to Tat with high affinity (300 nM). The characterization of a number of I?B-?-based peptides also provided insights into their intrinsic folding properties. These findings have been corroborated by mutagenesis studies on the full-length I?B-?, which unveil that different I?B-? residues are involved in NF-?B or Tat recognition.