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. Author manuscript; available in PMC: 2006 Jun 5.

Published in final edited form as: Biochemistry. 2003 Mar 11;42(9):2634–2642. doi: 10.1021/bi0271190

Influence of nucleocapsid protein on the dimerization of 1-444 and 1-561 RNAs. (A) HIV-1 Lai 1-290 RNA was incubated for 30 minutes at 37° or 55°C with or without HIV-1 nucleocapsid protein NCp7 in monomer buffer (50 mM Tris-HCl pH7.5, 40 mM potassium and 0.1 mM magnesium). After SDS/phenol extraction, the samples were loaded on a TBE/24°C gel. (B) HIV-2 ROD 1-444 (left panel) and 1-561 (right panel) RNAs were incubated as in (A). Two supplementary lanes showed incubation of the RNAs in dimer buffer (high potassium/high magnesium) without NCp7 at 37°C (lanes 7 and 15) or 55°C (lanes 8 and 16). SDS/phenol extracted samples are boxed. (C) 1-444 wild type (lanes 1-3) or 1-444 with the GUAC sequence deleted from the apical SL1 loop (lanes 4-6) were dimerized at 37°C with or without NCp, or at 55°C without NCp, as indicated. The RNAs lacking an intact SL1 were incapable of forming a TBE stable tight dimer. (D) A plot of dimerization yield as a function of SDS/phenol extraction and presence of NCp7 at 37°C for the 1-444 (open bars) and 1-561 (filled bars) RNAs. The y-axis error bars represent the standard deviation of two experiments.

Influence of nucleocapsid protein on the dimerization of 1-444 and 1-561 RNAs. (A) HIV-1 Lai 1-290 RNA was incubated for 30 minutes at 37° or 55°C with or without HIV-1 nucleocapsid protein NCp7 in monomer buffer (50 mM Tris-HCl pH7.5, 40 mM potassium and 0.1 mM magnesium). After SDS/phenol extraction, the samples were loaded on a TBE/24°C gel. (B) HIV-2 ROD 1-444 (left panel) and 1-561 (right panel) RNAs were incubated as in (A). Two supplementary lanes showed incubation of the RNAs in dimer buffer (high potassium/high magnesium) without NCp7 at 37°C (lanes 7 and 15) or 55°C (lanes 8 and 16). SDS/phenol extracted samples are boxed. (C) 1-444 wild type (lanes 1-3) or 1-444 with the GUAC sequence deleted from the apical SL1 loop (lanes 4-6) were dimerized at 37°C with or without NCp, or at 55°C without NCp, as indicated. The RNAs lacking an intact SL1 were incapable of forming a TBE stable tight dimer. (D) A plot of dimerization yield as a function of SDS/phenol extraction and presence of NCp7 at 37°C for the 1-444 (open bars) and 1-561 (filled bars) RNAs. The y-axis error bars represent the standard deviation of two experiments.