NCp9 are signatures of a ‘slow’ DNA binding protein. Despite this slow binding characteristic, the capability of Gag to aggregate nucleic acids, most likely makes it a reasonably good chaperone for processes that do not require signiﬁcant duplex destabilization, such as tRNA annealing (51–53). However, processes that require signiﬁcant duplex destabil- ization, such as strand transfer, will likely be optimally facil- itated by the ﬁnal processed NCp7, which exhibits strong duplex destabilization, and which facilitates rapid reannealing of DNA that has been melted by force. The direct results of the studies presented here can be sum- marized as follows. Both single DNA molecule stretching experiments and bulk Trp ﬂuorescence assays showed that the three Gag cleavage products NCp15, NCp9 and NCp7 bound DNA with comparable afﬁnity. The FA assays showed that NC and Gag bind with similar afﬁnity to short DNA and RNA oligonucleotides. In contrast, the DNA stretching experi- ments showed that Gag bound single polymeric DNA molecules with an apparent afﬁnity 10 times higher than that of any of the Gag cleavage products. Based on the change in single DNA molecule melting force observed in the pres- ence of these proteins, we also found that Gag was unable to destabilize the DNA helix, while the Gag cleavage products destabilized DNA signiﬁcantly. Finally, upon relaxation of the DNA after force-induced melting, we found that the amount of hysteresis was greatest for Gag, followed by NCp9 and NCp15. The ﬁnal NC Gag cleavage product, NCp7, exhibited the least amount of hysteresis, indicating that this protein is most efﬁcient at reannealing the regions of a single DNA molecule that have been melted by force. Thus, the ﬁnal Gag cleavage product is expected to be most efﬁcient nucleic acid chaperone for processes that require both nucleic acid destabilization and nucleic acid annealing. Both of these characteristics are required for minus-strand transfer, in which the very stable nucleic acid secondary structures, TAR RNA and cTAR DNA, must be annealed to each other. In contrast, Gag is more efﬁcient at binding and packaging viral RNA. It can also perform some nucleic acid annealing functions, such as tRNA annealing, a process that does not require signiﬁcant duplex destabilization, but involves nucleic acid aggregation by protein. ACKNOWLEDGEMENTS This work was funded in part by the National Cancer Institute under contract No. NO1-CO-12400 with SAIC-Frederick, Inc., by the National Science Foundation (MCB-0238190), the National Institutes of Health (GM-072462), by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research, and the Research Corporation. 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