PMID- 10526235
OWN - NLM
STAT- MEDLINE
DCOM- 19991223
LR  - 20071115
IS  - 1356-9597 (Print)
IS  - 1356-9597 (Linking)
VI  - 4
IP  - 8
DP  - 1999 Aug
TI  - Two separate sequences of PB2 subunit constitute the RNA cap-binding site of
      influenza virus RNA polymerase.
PG  - 475-85
AB  - BACKGROUND: Influenza virus RNA polymerase with the subunit composition of
      PB1-PB2-PA is a unique multifunctional enzyme with the activities of both
      synthesis and cleavage of RNA, and is involved in both transcription and
      replication of the RNA genome. Transcription is initiated by using capped RNA
      fragments, which are generated after cleavage of host cell mRNA by the RNA
      polymerase-associated capped RNA endonuclease. To identify the RNA cap 1-binding 
      site on the RNA polymerase, viral ribonucleoprotein (RNP) cores were subjected to
      UV-crosslinking with RNA which was labelled with 32P only at the cap-1 structure.
      RESULTS: After SDS-PAGE of UV-crosslinked cores, 32P was found to be associated
      only with the PB2 subunit (759 amino acid residues). The labelled PB2 was
      subjected, together with PB2 expressed in E. coli, to limited digestion with V8
      protease. Analysis of the amino terminal sequences of some isolated fragments
      with the crosslinked cap-1 indicated that two separate sequences within the PB2
      were involved in RNA cap-1 binding, one (N-site) at the N-terminal proximal
      region approximately between amino acid residues 242-282 downstream from the PB1 
      subunit-binding site and the other (C-site) between residues 538-577 including
      the cap-binding motifs. Two lines of evidence support the prediction of the
      involvement of two separate PB2 sequences on the RNA cap-binding: (i)
      cross-linking of the capped RNA on to expressed and isolated PB2 fragments, each 
      containing either the N-site or the C-site; and (ii) competition of capped
      RNA-binding to PB2 by both of the N- and C-terminal PB2 fragments. Taking
      together, we propose that two separate sequences within PB2 constitute the capped
      RNA-binding site of the RNA polymerase. CONCLUSION: Two separate sequences, one
      N-(242-282) and the other C-terminal (538-577) proximal segments of PB2 subunit, 
      constitute the RNA cap-binding site of the influenza virus RNA polymerase.
FAU - Honda, A
AU  - Honda A
AD  - Department of Molecular Genetics, National Institute of Genetics, Mishima,
      Shizuoka 411-8540, Japan.
FAU - Mizumoto, K
AU  - Mizumoto K
FAU - Ishihama, A
AU  - Ishihama A
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - England
TA  - Genes Cells
JT  - Genes to cells : devoted to molecular & cellular mechanisms
JID - 9607379
RN  - 0 (Carrier Proteins)
RN  - 0 (PB2 protein, Influenzavirus A)
RN  - 0 (RNA Caps)
RN  - 0 (RNA, Viral)
RN  - 0 (TNF Receptor-Associated Factor 3)
RN  - 0 (TRAF3 protein, human)
RN  - 0 (Tumor Necrosis Factor Receptor-Associated Peptides and Proteins)
RN  - 0 (Viral Proteins)
RN  - EC 2.5.1.18 (Glutathione Transferase)
RN  - EC 2.7.7.48 (RNA Replicase)
RN  - EC 2.7.7.6 (DNA-Directed RNA Polymerases)
RN  - EC 3.1.- (Endonucleases)
SB  - IM
MH  - Carrier Proteins/genetics
MH  - DNA-Directed RNA Polymerases/*genetics
MH  - Endonucleases/metabolism
MH  - Glutathione Transferase/metabolism
MH  - Kinetics
MH  - Models, Genetic
MH  - Orthomyxoviridae/*genetics
MH  - *RNA Caps
MH  - RNA Replicase
MH  - RNA, Viral/genetics
MH  - TNF Receptor-Associated Factor 3
MH  - *Tumor Necrosis Factor Receptor-Associated Peptides and Proteins
MH  - Ultraviolet Rays
MH  - Viral Proteins/*genetics/metabolism
EDAT- 1999/10/20 00:00
MHDA- 1999/10/20 00:01
CRDT- 1999/10/20 00:00
PHST- 1999/10/20 00:00 [pubmed]
PHST- 1999/10/20 00:01 [medline]
PHST- 1999/10/20 00:00 [entrez]
AID - gtc275 [pii]
PST - ppublish
SO  - Genes Cells. 1999 Aug;4(8):475-85.