Analysis of the nucleic acid annealing activities of nucleocapsid protein from HIV-1.
Retroviral nucleocapsid (NC) protein is an integral part of the virion nucleocapsid where it is in tight association with genomic RNA and the tRNA primer. NC protein is necessary for the dimerization and encapsidation of genomic RNA, the annealing of the tRNA primer to the primer binding site (PBS) and the initial strand transfer event. Due to the general nature of NC protein-promoted annealing, its use to improve nucleic acid interactions in various reactions can be envisioned. Parameters affecting NC-promoted nucleic acid annealing of NCp7 from HIV-1 have been analyzed. The promotion of RNA:RNA and RNA:DNA annealing by NCp7 is more sensitive to the concentration of MgCl2 than the promotion of DNA:DNA hybridization. Stimulation of complex formation for all three complexes was efficient at 0-90 mM NaCl, between 23 and 55 degrees C and at pH values between 6.5 and 9.5, inclusive. Parameters affecting NCp7-promoted hybridization of tRNA(Lys,3) to the PBS, which appears to be specific for NC protein, will be discussed. Results implicate the basic regions of NCp7, but not the zinc fingers, in promoting the annealing of complementary nucleic acid sequences. Finally, NCp7 strand transfer activity aids the formation of the most stable nucleic acid complex.
Lapadat-Tapolsky
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M
Unité de Virologie Humaine (INSERM 412), LaboRétro, Ecole Normale Supérieure de Lyon, France.
Pernelle
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C
Borie
C
C
Darlix
J L
JL
D001483:Base Sequence; D002213:Capsid / Q000494:pharmacology*; D036022:Capsid Proteins; D004247:DNA / Q000378:metabolism*; D015683:Gene Products, gag / Q000494:pharmacology*; D015497:HIV-1 / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D015636:Magnesium Chloride / Q000494:pharmacology; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D012313:RNA / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D013696:Temperature; D014764:Viral Proteins; D016335:Zinc Fingers; D054301:gag Gene Products, Human Immunodeficiency Virus
D036022:Capsid Proteins; D015683:Gene Products, gag; C071884:NCP7 protein, Human immunodeficiency virus 1; D012346:RNA, Transfer, Amino Acyl; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; D015636:Magnesium Chloride; D012965:Sodium Chloride; D012313:RNA; D004247:DNA