title: Characterization of the archaeal ribonuclease P proteins from Pyrococcus horikoshii OT3.
abstract: Ribonuclease P (RNase P) is a ribonucleoprotein complex involved in the processing of the 5'-leader sequence of precursor tRNA (pre-tRNA). Our earlier study revealed that RNase P RNA (pRNA) and five proteins (PhoPop5, PhoRpp38, PhoRpp21, PhoRpp29, and PhoRpp30) in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 reconstituted RNase P activity that exhibits enzymatic properties like those of the authentic enzyme. In present study, we investigated involvement of the individual proteins in RNase P activity. Two particles (R-3Ps), in which pRNA was mixed with three proteins, PhoPop5, PhoRpp30, and PhoRpp38 or PhoPop5, PhoRpp30, and PhoRpp21 showed a detectable RNase P activity, and five reconstituted particles (R-4Ps) composed of pRNA and four proteins exhibited RNase P activity, albeit at reduced level compared to that of the reconstituted particle (R-5P) composed of pRNA and five proteins. Time-course analysis of the RNase P activities of R-4Ps indicated that the R-4Ps lacking PhoPop5, PhoRpp21, or PhoRpp30 had virtually reduced activity, while omission of PhoRpp29 or PhoRpp38 had a slight effect on the activity. The results indicate that the proteins contribute to RNase P activity in order of PhoPop5 > PhoRpp30 > PhoRpp21 >> PhoRpp29 > PhoRpp38. It was further found that R-4Ps showed a characteristic Mg2+ ion dependency approximately identical to that of R-5P. However, R-4Ps had optimum temperature of around at 55 degrees C which is lower than 70 degrees C for R-5P. Together, it is suggested that the P. horikoshii RNase P proteins are predominantly involved in optimization of the pRNA conformation, though they are individually dispensable for RNase P activity in vitro.
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School, Kyushu University, Hakozaki 6-10-1, Higashi-ku, Fukuoka 812-8581.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Terada
authors.forename: Takashi
authors.initials: T
authors.lastname: Honda
authors.forename: Hideo
authors.initials: H
authors.lastname: Fukuhara
authors.forename: Kazumasa
authors.initials: K
authors.lastname: Hada
authors.forename: Makoto
authors.initials: M
authors.lastname: Kimura
mesh_terms: D019843:Archaeal Proteins / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D021122:Protein Subunits; D043902:Pyrococcus horikoshii / Q000378:metabolism*; D016337:RNA, Catalytic / Q000737:chemistry / Q000378:metabolism; D043262:Ribonuclease P / Q000737:chemistry / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; D021122:Protein Subunits; D016337:RNA, Catalytic; D043262:Ribonuclease P