ИСТИНА

Ku70/Ku80 is a heterodimer complex displaying essential functions for eukaryotic cell survival. Recently Ku has been identified as a co-factor for HIV-1 integrase (IN), which protects it from proteasomal degradation. In/Ku70 complex appears to be a promising drug target. The drug design would be facilitated if a detailed structure of Ku70/IN complex were available. The exact structure of HIV-1 integrase is not yet known, and only single domains of IN can be effectively crystallized. To learn the structure of Ku70/IN complex, we performed a systematic analysis of subdomains within IN that are required for the complex formation. Earlier by pull-down assay, we have shown that IN and Ku70 form a stable complex with a Kd about 70 nM. Here the use of the same technique and SPR-analysis allowed us to elucidate interactions of full-length Ku70 with various IN fragments. N-His6-tagged HIV-1 IN individual domains [N-terminal (1– 50 aa), catalytic (51–220 aa) and C-terminal (220–270 aa)] and several truncated variants containing amino acids 1–160, 1–220, 51–160, 51–220 and 51–280 were expressed in E. coli. A full-size Ku70 with an N-terminal GST-tag was also expressed in E. coli. Our data shows that neither N-terminal nor C-terminal domains of IN are essential for its binding to Ku70. The affinity to Ku70 of the catalytic core (51–220 aa) was comparable to the affinity of the full-size IN, whereas its truncated variant (51–160 aa) bound to Ku70 only weakly. Thus, the region of IN from 160 to 220 aa is extremely important for Ku70/IN complex formation.