Protein translocation across the membrane, aided by specific receptors, is a ubiquitous and essential process in every cell. The receptors involved in protein translocation can be classified into two categories: 1) those that recognize precursor proteins through interaction with their specific pre-sequences and 2) those that recognize cytosolic transporters (chaperones) which are involved in the delivery of precursor proteins to the import machinery for the specific organelle to which they are targeted [1,2]. In the latter case, often the Tetratricopeptide Repeat (TPR) motifs are present in receptors and they have been shown to aid in the recognition of precursor protein pre-sequences. The TPR motif therefore, plays an important function in its ability to form protein-protein interactions.

The Translocon outer membrane chloroplast 64kDa (Toc64) protein is a component of Toc core protein complex in Arabidopsis thaliana (At) [3]. It has a C-terminal 3-TPR domain, which is hypothesized to recognize the conserved C-terminus of the chaperone proteins, Hsp70 and Hsp90. In the present study, we focus on the energetics and molecular basis of C-term Hsp70 and C-term Hsp90 interaction with AtToc64. Here we have quantitatively assessed the contribution of TPR-mediated peptide recognition to complex formation using Isothermal Titration Calorimetry. In addition, in silico simulation studies, combined with empirical molecular mechanics have been undertaken to understand the detailed interaction profile. Guided by the above methods, we are able to identify important residues in the ligands that mediate interaction with the TPR domain of AtToc64.