We have studied the conformational dynamics associated with the nanoscale DNA bending induced by human immunodeficiency virus type 1 (HIV-1) nucleocapsid (NC) protein using single-molecule Forster resonance energy transfer (SM-FRET). To gain molecular-level insights into how the HIV-1 NC locally distorts the structures of duplexed DNA segments, the dynamics, reversibility, and sequence specificity of the DNA bending behavior of NC have been systematically studied. We have performed SM-FRET measurements on a series of duplexed DNA segments with varying sequences, lengths, and local structures in the presence of the wide-type HIV-1 NC and NC mutants lacking either the basic N-terminal domain or the zinc fingers. On the basis of the SM-FRET results, we have proposed a possible mechanism for the NC-induced DNA bending in which both NC's zinc fingers and N-terminal domain are found to play crucial roles. The SM-FRET results reported here add new mechanistic insights into the biological behaviors and functions of HIV-1 NC as a retroviral DNA-architectural protein which may play critical roles in the compaction, nuclear import, and integration of the proviral DNA during the retroviral life cycle.