HIV-1 nucleocapsid protein NCp7 and its RNA stem loop 3 partner: rotational dynamics of spin-labeled RNA stem loop 3

Biochemistry
Xiangmei XiCharles P Scholes

Abstract

The tumbling dynamics of a 20-mer HIV-1 RNA stem loop 3 spin-labeled at the 5' position were probed in the nanosecond time range. This RNA interacted with the HIV-1 nucleocapsid Zn-finger protein, 1-55 NCp7, and specialized stopped-flow EPR revealed concomitant kinetics of probe immobilization from milliseconds to seconds. RNA stem loop 3 is highly conserved in HIV, while NCp7 is critical to HIV-RNA packaging and annealing. The 5' probe did not perturb RNA melting or the NCp7/RNA interaction monitored by gel shift and fluorescence. The 5'-labeled RNA tumbled with a subnanosecond isotropic correlation time (approximately 0.60 ns at room temperature) reflecting both local viscosity-independent bond rotation of the probe and viscosity-dependent diffusion of 40-60% of the RNA. The binding of NCp7 to spin-labeled RNA stem loop 3 in a 1:1 ratio increased the spin-labeled tumbling time by about 40%. At low ionic strength with a ratio of NCp7 to RNA >or=3 (i.e., an NCp7 to nucleotide ratio <or=7, which is the threshold ratio for chaperone effects), the probe tumbling time markedly increased to several nanoseconds, signifying a NCP7/RNA complex with restricted motion even at the initially mobile 5' position. Increasing the ionic strengt...Continue Reading

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Citations

Jul 19, 2011·Journal of Molecular Biology·Jean-Luc DarlixYves Mély
Feb 19, 2015·Biophysical Journal·Yan SunCharles P Scholes
Sep 2, 2011·Wiley Interdisciplinary Reviews. RNA·Phuong Nguyen, Peter Z Qin
Mar 6, 2010·Journal of Chemical Information and Modeling·Mattia MoriMaurizio Botta

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