C-terminal domain modulates the nucleic acid chaperone activity of human T-cell leukemia virus type 1 nucleocapsid protein via an electrostatic mechanism.
Retroviral nucleocapsid (NC) proteins are molecular chaperones that facilitate nucleic acid (NA) remodeling events critical in viral replication processes such as reverse transcription. Surprisingly, the NC protein from human T-cell leukemia virus type 1 (HTLV-1) is an extremely poor NA chaperone. Using bulk and single molecule methods, we find that removal of the anionic C-terminal domain (CTD) of HTLV-1 NC results in a protein with chaperone properties comparable with that of other retroviral NCs. Increasing the ionic strength of the solution also improves the chaperone activity of full-length HTLV-1 NC. To determine how the CTD negatively modulates the chaperone activity of HTLV-1 NC, we quantified the thermodynamics and kinetics of wild-type and mutant HTLV-1 NC/NA interactions. The wild-type protein exhibits very slow dissociation kinetics, and removal of the CTD or mutations that eliminate acidic residues dramatically increase the protein/DNA interaction kinetics. Taken together, these results suggest that the anionic CTD interacts with the cationic N-terminal domain intramolecularly when HTLV-1 NC is not bound to nucleic acids, and similar interactions occur between neighboring molecules when NC is NA-bound. The intramolecular N-terminal domain-CTD attraction slows down the association of the HTLV-1 NC with NA, whereas the intermolecular interaction leads to multimerization of HTLV-1 NC on the NA. The latter inhibits both NA/NC aggregation and rapid protein dissociation from single-stranded DNA. These features make HTLV-1 NC a poor NA chaperone, despite its robust duplex destabilizing capability.
Qualley
Dominic F
DF
Department of Chemistry and Biochemistry, Center for Retrovirus Research, Ohio State University, Columbus, Ohio 43210, USA.
Stewart-Maynard
Kristen M
KM
Wang
Fei
F
Mitra
Mithun
M
Gorelick
Robert J
RJ
Rouzina
Ioulia
I
Williams
Mark C
MC
Musier-Forsyth
Karin
K
D000595:Amino Acid Sequence; D001483:Base Sequence; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D015368:Human T-lymphotropic virus 1 / Q000737:chemistry*; D007700:Kinetics; D018832:Molecular Chaperones / Q000378:metabolism*; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation / Q000187:drug effects; D009691:Nucleic Acid Denaturation / Q000187:drug effects; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry; D009696:Nucleic Acids / Q000378:metabolism*; D019590:Nucleocapsid Proteins / Q000737:chemistry; D009994:Osmolar Concentration; D017434:Protein Structure, Tertiary; D012367:RNA, Viral / Q000737:chemistry; D012965:Sodium Chloride / Q000494:pharmacology; D012996:Solutions; D055672:Static Electricity; D016335:Zinc Fingers
D004279:DNA, Viral; D018832:Molecular Chaperones; D050505:Mutant Proteins; D009692:Nucleic Acid Heteroduplexes; D009696:Nucleic Acids; D019590:Nucleocapsid Proteins; D012367:RNA, Viral; D012996:Solutions; D012965:Sodium Chloride