Histone-binding domains in a human nuclear autoantigenic sperm protein.

In one of our previous studies, the deduced amino acid sequence of the human nuclear autoantigenic sperm protein (hNASP) revealed two conserved histone-binding domains when compared to the Xenopus N1/N2 protein sequence. These histone-binding domains of Xenopus N1/N2 are known to be functional; however, their function in hNASP is unknown. In this study we have determined the number, location, and activity of the histone-binding domains on the primary sequence of hNASP. Purified recombinant polypeptides expressing the full-length hNASP and various deletion constructs covering the entire length of the hNASP sequence were tested by Western blotting and in ELISA for binding to biotin-labeled histones. A positive reaction was detected for the full-length recombinant protein and for the polypeptides spanning the N-terminal region (amino acids [aa] 32-192), and two additional regions: aa 193-352 and aa 353-572. The lack of binding to the expressed C-terminal (aa 573-787), which also contains polyacidic amino acids, suggests that the binding of hNASP to the somatic core histones is a sequence-specific as well as an electrostatic interaction. The removal of flanking sequences from the binding domains did not abrogate their ability to bind histones. We conclude that there are at least three functional histone-binding domains in hNASP, two of them encompassing the predicted histone binding sites homologous to the N1/N2 protein, and a third novel domain. Therefore, hNASP may be defined as a nuclear histone-binding protein found in human testis.