Multimerization of HIV-1 integrase hinges on conserved SH3-docking platforms

Meytal Galilee; Akram Alian

The higher number of interactions and the larger surface area buried upon peptide binding to site-2 may highlight a binding privilege at this site. Analyses of crystal structures of IN truncation variants, which have been shown to dimerize (CCD and CCDCTD) or tetramerize (NTDCCD) in solution and crystals, show the clear exposure and accessibility of site-1 ( Figure 3A and B). Whereas site-2 is also exposed in the crystal structures of dimeric CCD and tetrameric NTDCCD ( Figure 3A ), analyses of CCDCTD crystal structures intriguingly show that CTD is constantly docked into site-2, albeit in various configurations distinct to that of the functional intasome ( Figure 3B , orange cartoon). If indeed the CTD preoccupies site-2 within the truncated CCDCTD variant, as seen in crystal structures, then we expect that, unlike CCD or NTDCCD variants, the CCDCTD construct would not bind the peptide. Assessing peptide-binding affinities to the various IN constructs show that the presence of NTD (within NTDCCD) only slightly (~ 1.5 folds) interferes with peptide binding ( Figure 3C ). CTD (within CCDCTD), on the other hand, almost completely abolishes peptide binding to more than 12 folds ( Figure 3C ). In agreement with the binding data, structural superimposition of the peptide shows that while NTD (of NTDCCD structure) may delicately interfere with peptide binding at site-1 ( Figure 3A ), CTD (of CCDCTD structures) would provide a more robust interference at site-2 ( Figure 3B ). It is worth noting that the peptide bound to full-length IN, which is preassembled into tetramers in solution (24) , with affinities (Kd 1700 ± 88 nM) corresponding to the concentrations in the monomerization assay ( Figure 1E ) and ~ 4 folds weaker than NTDCCD (Kd 441 ± 11 nM, Figure 3C ). Together, the NTD appears to crucially modulate CTD interactions with the CCD.