title: Functional implication of archaeal homologues of human RNase P protein pair Pop5 and Rpp30.
abstract: PhoPop5 and PhoRpp30 in the hyperthermophilic archaeon Pyrococcus horikoshii, homologues of human ribonuclease P (RNase P) proteins hPop5 and Rpp30, respectively, fold into a heterotetramer [PhoRpp30-(PhoPop5)2-PhoRpp30], which plays a crucial role in the activation of RNase P RNA (PhopRNA). Here, we examined the functional implication of PhoPop5 and PhoRpp30 in the tetramer. Surface plasmon resonance (SPR) analysis revealed that the tetramer strongly interacts with an oligonucleotide including the nucleotide sequence of a stem-loop SL3 in PhopRNA. In contrast, PhoPop5 had markedly reduced affinity to SL3, whereas PhoRpp30 had little affinity to SL3. SPR studies of PhoPop5 mutants further revealed that the C-terminal helix (α4) in PhoPop5 functions as a molecular recognition element for SL3. Moreover, gel filtration indicated that PhoRpp30 exists as a monomer, whereas PhoPop5 is an oligomer in solution, suggesting that PhoRpp30 assists PhoPop5 in attaining a functionally active conformation by shielding hydrophobic surfaces of PhoPop5. These results, together with available data, allow us to generate a structural and mechanistic model for the PhopRNA activation by PhoPop5 and PhoRpp30, in which the two C-terminal helices (α4) of PhoPop5 in the tetramer whose formation is assisted by PhoRpp30 act as binding elements and bridge SL3 and SL16 in PhopRNA.
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Graduate School, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan and.
authors.forename: Masato
authors.initials: M
authors.lastname: Hamasaki
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Graduate School, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan and.
authors.forename: Kohsuke
authors.initials: K
authors.lastname: Hazeyama
authors.affiliation: Laboratory of Structural Biology, Division of Bioengineering, Graduate School of Systems Life Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan.
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Iwasaki
authors.affiliation: Laboratory of Structural Biology, Division of Bioengineering, Graduate School of Systems Life Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan.
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Ueda
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Graduate School, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan and Laboratory of Structural Biology, Division of Bioengineering, Graduate School of Systems Life Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Nakashima
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Graduate School, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan and Laboratory of Structural Biology, Division of Bioengineering, Graduate School of Systems Life Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan.
authors.forename: Yoshimitsu
authors.initials: Y
authors.lastname: Kakuta
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Graduate School, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan and Laboratory of Structural Biology, Division of Bioengineering, Graduate School of Systems Life Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan mkimura@agr.kyushu-u.ac.jp.
authors.forename: Makoto
authors.initials: M
authors.lastname: Kimura
mesh_terms: D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D001324:Autoantigens / Q000235:genetics; D001483:Base Sequence; D006801:Humans; D007700:Kinetics; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D043902:Pyrococcus horikoshii / Q000235:genetics*; D019642:RNA, Archaeal / Q000378:metabolism*; D043262:Ribonuclease P / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D020349:Surface Plasmon Resonance
chemical_list: D019843:Archaeal Proteins; D001324:Autoantigens; D018832:Molecular Chaperones; D021122:Protein Subunits; D019642:RNA, Archaeal; C104646:RPP30 protein, human; C000605944:Pop5 protein, human; D043262:Ribonuclease P
keywords: Pyrococcus horikoshii; archaea; protein–RNA interaction; ribonuclease P; surface plasmon resonance