Studies on Methanocaldococcus jannaschii RNase P reveal insights into the roles of RNA and protein cofactors in RNase P catalysis.
Ribonuclease P (RNase P), a ribonucleoprotein (RNP) complex required for tRNA maturation, comprises one essential RNA (RPR) and protein subunits (RPPs) numbering one in bacteria, and at least four in archaea and nine in eukarya. While the bacterial RPR is catalytically active in vitro, only select euryarchaeal and eukaryal RPRs are weakly active despite secondary structure similarity and conservation of nucleotide identity in their putative catalytic core. Such a decreased archaeal/eukaryal RPR function might imply that their cognate RPPs provide the functional groups that make up the active site. However, substrate-binding defects might mask the ability of some of these RPRs, such as that from the archaeon Methanocaldococcus jannaschii (Mja), to catalyze precursor tRNA (ptRNA) processing. To test this hypothesis, we constructed a ptRNA-Mja RPR conjugate and found that indeed it self-cleaves efficiently (k(obs), 0.15 min(-1) at pH 5.5 and 55 degrees C). Moreover, one pair of Mja RPPs (POP5-RPP30) enhanced k(obs) for the RPR-catalyzed self-processing by approximately 100-fold while the other pair (RPP21-RPP29) had no effect; both binary RPP complexes significantly reduced the monovalent and divalent ionic requirement. Our results suggest a common RNA-mediated catalytic mechanism in all RNase P and help uncover parallels in RNase P catalysis hidden by plurality in its subunit make-up.
Pulukkunat
Dileep K
DK
Ohio State Biochemistry Program and Department of Biochemistry, The Ohio State University, Columbus, OH 43210, USA.
Gopalan
Venkat
V
D019843:Archaeal Proteins / Q000737:chemistry* / Q000378:metabolism; D002384:Catalysis; D007700:Kinetics; D017015:Methanococcales / Q000201:enzymology*; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D012322:RNA Precursors / Q000737:chemistry / Q000378:metabolism; D019642:RNA, Archaeal / Q000737:chemistry* / Q000378:metabolism; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism; D012365:RNA, Transfer, Tyr / Q000737:chemistry / Q000378:metabolism; D043262:Ribonuclease P / Q000737:chemistry* / Q000378:metabolism
D019843:Archaeal Proteins; D021122:Protein Subunits; D012322:RNA Precursors; D019642:RNA, Archaeal; D012365:RNA, Transfer, Tyr; D012343:RNA, Transfer; D043262:Ribonuclease P