Analysis of DNA compaction profile and intracellular contents of archaeal histones from Pyrococcus kodakaraensis KOD1.
Two histone genes, hpkA and hpkB, from hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1 strain were cloned, sequenced, and expressed in Escherichia coli cells. Both hpkA and hpkB genes encoded a protein of 67 amino acids, however they possessed the different molecular weight (HpkA, 7,378:HpkB, 7,167). Deduced amino acid sequences of HpkA and HpkB were homologous to other archaeal histones and eucaryal core histones (H2A, H4). Gel mobility shift assays by purified proteins demonstrated that HpkB possessed higher affinity to DNA and more extensive ability to compact DNA than HpkA. HpkB prevented double stranded DNA from thermal denaturation in less amount than HpkA in vitro. In order to investigate intracellular contents of HpkA and HpkB in KOD1 cells, immunoblot analysis was performed by using anti-HpkA antisera obtained from immunized BALB/c mice, showing that HpkA was less abundantly expressed than HpkB in KOD1 cells. These results suggest that HpkB plays a major role to protect double stranded DNA from thermal denaturation in vivo.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002942:Circular Dichroism; D019641:DNA, Archaeal / Q000737:chemistry / Q000235:genetics*; D006657:Histones / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D006358:Hot Temperature; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009691:Nucleic Acid Denaturation; D011489:Protein Denaturation; D019711:Pyrococcus / Q000235:genetics*; D017386:Sequence Homology, Amino Acid