Human T-Cell Lymphotropic Virus Type 1 Nucleocapsid Protein-Induced Structural Changes in Transactivation Response DNA Hairpin Measured by Single-Molecule Fluorescence Resonance Energy Transfer

Role of Conformational Dynamics in α-Amino-3-hydroxy-5-methylisoxazole-4-propionic Acid (AMPA) Receptor Partial Agonism

“…The sites tagged with the donor and acceptor fluorophores are T394C and S652C, similar to those used in our previous smFRET investigations (30). The smFRET data from a number of such traces (170 -210 traces) for each willardiine were then processed for background and cross-talk correction (35,36) and denoised using wavelet decomposition as described previously (37). The data were then plotted as histograms of fraction of occurrence versus the FRET efficiency to determine the spread of states that the protein explores (Fig.…”

“…Experimental Setup for smFRET-All single-molecule fluorescence measurements were performed using a custom built confocal microscope (35,36). A 532-nm diode-pumped solid state laser (Coherent, Compass 315M-100 SL) was used for sample excitation.…”

Retrovirus-Specific Differences in Matrix and Nucleocapsid Protein-Nucleic Acid Interactions: Implications for Genomic RNA Packaging

“…With a 228-nt gRNA R region, which is predicted to fold into a complex secondary structure (57), HTLV-1 would be expected to require a robust chaperone to facilitate the minus-strand transfer steps of reverse transcription. Surprisingly, HTLV-1 and -2 NC proteins display relatively poor NA binding and overall chaperone activity; however, once bound, HTLV-1 NC is a strong duplex destabilizer (28,29). We therefore suggest that in deltaretroviruses, MA is involved in key early steps involving genome recognition and packaging; however, once the local concentration of Gag is high enough, MA preferentially binds to the membrane, and NC domain binding to NAs occurs, allowing NC to carry out the chaperone function required for reverse transcription (Fig.…”

“…Interestingly, HTLV-1 NC displays reduced NA binding affinity and chaperone function relative to those of HIV-1 NC yet has robust duplex-destabilizing capabilities (28,29). Our laboratory has previously explored the mechanistic basis for the poor NA binding and chaperone properties of HTLV-1 NC, and our studies show that removal of HTLV-1 NC's anionic C-terminal domain (CTD) improves the chaperone function to a level comparable to those of other retroviral NCs (30).…”

Single-molecule stretching studies of RNA chaperones

“…95 At the same time, these NC proteins' ability to destabilize DNA duplex appeared to be rather similar for all tested NC proteins, including the worst chaperone-HTLV-1 NC. [95][96][97] Although some defects in the ability of mutant NC proteins to facilitate NA annealing were consistently observed, 67,92,95,96,98,99 these defects are not nearly as strong as the up to 6 orders of magnitude decrease in viral infectivity measured in vivo for the viruses with the same mutations in NC. [100][101][102] While simple sedimentation assays suggest that these mutant NC proteins are still capable of aggregating NA, 41,51,95,96 it is possible that these "slow" NC proteins may interfere with reverse transcription by partially losing their ability to tightly aggregate NA within the capsid, thereby leading to early capsid uncoating prior to the completion of the RTion.…”