Domain Analysis of the Chloroplast Polynucleotide Phosphorylase Reveals Discrete Functions in RNA Degradation, Polyadenylation, and Sequence Homology with Exosome Proteins. The Plant Cell , Vol. 15, 2003-2019, September 2003
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Domain Analysis of the Chloroplast Polynucleotide Phosphorylase Reveals Discrete Functions in RNA Degradation, Polyadenylation, and Sequence Homology with Exosome Proteins
The Plant Cell, Vol. 15, 2003-2019, September 2003
Shlomit Yehudai-Resheffa, Victoria Portnoya, Sivan Yogeva, Noam Adirb and Gadi Schuster1
1.chloroplast mRNA degradation (chloroplast and bacteria)
1). endonucleolytic cleavage
2). addition of poly(A)-rich sequences to the endonucleolytic cleavage products
(PAP in E.coli)
3). exonucleolytic degradation
2.PNPase in the chloroplast was found to form a homotrimeric complex
and lacks any known interactions with other proteins.
RNA. 7, (2001), 1464–1475
3.No PAP can be detected in spinach chloroplasts, and thus both
polyadenylation and degradation are performed by one enzyme,
Mol. Cell. Biol.21, (2001), 5408–5416
core = RNase PH domain
The amino acid sequence and domain structure is largely conserved between bacteria and organelles.
# bacteria : domain 2 have activity only
Degradation : domain 1 and 2
Polymerization : domain 2
Product : NDP (TLC)
UV light cross-linking assay
(Lisitsky et al., 1997b; Lisitsky and Schuster, 1999).
UV light cross-linking competition assay
substrate : 32P-psbA RNA
domain 2 (only)
PNPase enzyme is its pausing at a stem-loop structure when processively degrading RNA.
EMBO J. (1996) 15, 1132–1141
E. coli PAP I is inhibited by a stem-loop structure but that the addition of two nucleotides 3' to the stem loop is sufficient to promote efficient polyadenylation.
Nucleic Acids Res.(2000) 28, 1139–1144.
E. coli strain SK 8992
E. ColiArcheal Yeast Human
PNPaseRrp41Rrp41p/Ski6phRrp41pRNase PH Q17533
(x 3)(x 3)Rrp46phRrp46pRNase PH (Crn-5)
Rrp42Rrp42phRrp42pRNase PH NP_508024
(x 3)Rrp43pOIP2RNase PH
Rrp45pPM/Scl-75RNase PH T28842
Other common proteins:
(RNase R?)Rrp44p/Dis3p (hDis3p)RNase R
(RNase D?)Rrp6pPM/Scl-100RNase D (Crn-3)(nuclear only)
PNPase (x3 ?)
red: in vitro 3’-5’exonuclease activity
PNPase lacks any known interactions with other proteins.