Structural Basis of Bispecific Protein:Protein Interactions

Abstract

The ability of the Escherichia coli protein, BirA, to act as both an enzyme and a transcription repressor allows communication between metabolism and gene expression. BirA forms a hetero-dimer for the essential post-translational biotin addition to acetyl-CoA carboxylase and forms a homo-dimer to bind site-specifically to DNA and repress transcription initiation at the biotin biosynthetic operon. A single surface on BirA is used for both protein:protein interactions. However, the extent to which the structural information on this surface is functionally shared by the two interactions is not known. Multiple loops on the surface are located in both dimer interfaces. In this work alanine substitution variants of two loops, composed of residues 140-146 and 193-199, have been constructed and the proteins have been purified. Measurements of homo- and hetero-dimerization energetics of loop variant proteins using sedimentation equilibrium indicate that several residues in the two loops contribute to both interactions. Thus, structural information in both loops was important for evolution of bifunctionality in BirA.Supported by NIH grants R01-GM46511 and S10-RR15899