Nucleobase-Specific Recognition of DNA by Protamine

Protamines--arginine (Arg)-rich, intrinsically disordered proteins (IDPs)--replace histones to compact DNA in the sperm nucleus. While folded domains can bind DNA site-specifically, whether IDPs exhibit DNA-sequence specificity remains open. Here, we used all-atom molecular dynamics simulations to characterize protamine-DNA binding. Eight copies of salmon protamine (33 residues with 21 Arg) rapidly wrap around a 35-bp DNA molecule. In addition to salt bridges with DNA phosphates, Arg hydrogen-bonds with multiple nucleobases on both strands or the same strand. At the major groove, protamine prefers GC over AT by a 2.6-to-1 ratio, with guanine's O6 and N7 atoms as hydrogen-bond acceptors; the preference is especially strong at G-tracts. This selectivity reverses at the minor groove, where AT overtakes GC by a 2.1-to-1 margin. The net effect is a moderate bias for GC over AT in protamine-base interactions. Additional simulations with 20-bp DNA molecules and free-energy calculations reinforce these observations. For experimental validation, we designed DNA sequences with 0-40% GC content. Protamine-DNA mixtures form aggregates, with an increasing propensity at higher GC content and upon shuffling into G-tracts. This work demonstrates that IDPs can bind DNA in a nucleobase-specific manner, with implications for protamine-mediated transcription silencing and remodeling of paternal chromatin.