A model of PSI dimerization: destabilization of the C278-G303 stem-loop by the nucleocapsid protein (NCp10) of MoMuLV.
We have shown that at low ionic strength (i.e., 100 mM NaCl) a short autocomplementary sequence spanning nucleotides C283 to G298 of MoMuLV RNA genome is involved in the process of PSI dimerization in vitro [Girard, P.-M., Bonnet-Mathonière, B., Muriaux, D., & Paoletti, J. (1995) Biochemistry 34, 9785-9794]. In order to identify other contributions of the PSI structure to RNA dimerization, we studied the kinetics of dimerization as a function of salt concentration of short RNA transcripts comprising or not the autocomplementary sequence C283-G298. We propose that, apart from the crucial role of this sequence in RNA dimerization, the 364-565 domain of PSI can interfere, in vitro, with the initiation of dimer formation. Intermolecular loop-loop recognitions involving the 364-565 domain could stabilize, in a salt concentration-dependent manner, a transient RNA dimer built around the loop-loop U288-A293 interaction. This dimer evolves toward a more stable structure which mainly corresponds to the annealing of two C283-G298 sequences. We also show that chemically synthesized NCp10 does not modify these steps but rather helps the system to pass over the energy barriers associated with the transition to stable RNA structures comprising the stem-loop C278-G303. Data obtained in the presence of NCp10 suggest a binding site size of 9 +/- 1 nucleotides per protein at 37 degrees C and a 10-20-fold increase in the rate constant (i.e., k1 = 24 000 +/- 7000 M-1 s-1) of dimer formation.
Girard
P M
PM
Unité de Biochimie, URA 147 CNRS, Institut Gustave Roussy, Villejuif, France.
de Rocquigny
H
H
Roques
B P
BP
Paoletti
J
J
D001483:Base Sequence; D015683:Gene Products, gag / Q000737:chemistry*; D007700:Kinetics; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000737:chemistry* / Q000235:genetics; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000494:pharmacology; D009994:Osmolar Concentration; D012367:RNA, Viral / Q000737:chemistry* / Q000187:drug effects; D012965:Sodium Chloride; D013696:Temperature; D014758:Viral Core Proteins / Q000737:chemistry*
D015683:Gene Products, gag; D009841:Oligonucleotides; D012367:RNA, Viral; D014758:Viral Core Proteins; C068662:retroviral nucleocapsid protein NCp10, Moloney murine leukemia virus; D012965:Sodium Chloride