PMID- 10531319
OWN - NLM
STAT- MEDLINE
DCOM- 19991216
LR  - 20210209
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 274
IP  - 44
DP  - 1999 Oct 29
TI  - Highly specific recognition of primer RNA structures for 2'-OH priming reaction 
      by bacterial reverse transcriptases.
PG  - 31236-44
AB  - A minor population of Escherichia coli contains retro-elements called retrons, 
      which encode reverse transcriptases (RT) to synthesize peculiar satellite DNAs 
      called multicopy single-stranded DNA (msDNA). These RTs recognize specific RNA 
      structures in their individual primer-template RNAs to initiate cDNA synthesis 
      from the 2'-OH group of a specific internal G residue (branching G residue). The 
      resulting products (msDNA) consist of RNA and single-stranded DNA, sharing hardly 
      any sequence homology. Here, we investigated how RT-Ec86 recognizes the specific 
      RNA structure in its primer-template RNA. On the basis of structural comparison 
      with HIV-1 RT, domain exchanges were carried out between two E. coli RTs, RT-Ec86 
      and RT-Ec73. RT-Ec86 (320 residues) and RT-Ec73 (316 residues) share only 71 
      identical residues (22%). From the analysis of 10 such constructs, the C-terminal 
      91-residue sequence of RT-Ec86 was found to be essential for the recognition of 
      the unique stem-loop structure and the branching G residue in the primer-template 
      RNA for retron-Ec86. Using the SELEX (systematic evolution of ligands by 
      exponential enrichment) method with RT-Ec86 and primer RNAs containing random 
      sequences, the identical stem-loop structure (including the 3-U loop) to that 
      found in the retron-Ec86 primer-template RNA was enriched. In addition, the 
      highly conserved 4-base sequence (UAGC), including the branching G residue, was 
      also enriched. These results indicate that the highly diverse C-terminal region 
      recognizes specific stem-loop structures and the branching G residue located 
      upstream of the stem-loop structure. The present results with seemingly primitive 
      RNA-dependent DNA polymerases provide insight into the mechanisms for specific 
      protein RNA recognition.
FAU - Inouye, S
AU  - Inouye S
AD  - Department of Biochemistry, Robert Wood Johnson Medical School, Piscataway, New 
      Jersey 08854, USA.
FAU - Hsu, M Y
AU  - Hsu MY
FAU - Xu, A
AU  - Xu A
FAU - Inouye, M
AU  - Inouye M
LA  - eng
GR  - GM 44012/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (DNA, Single-Stranded)
RN  - 0 (Ligands)
RN  - 0 (RNA primers)
RN  - 0 (RNA, Bacterial)
RN  - 0 (Recombinant Fusion Proteins)
RN  - 0 (Retroelements)
RN  - 0 (msDNA)
RN  - 63231-63-0 (RNA)
RN  - EC 2.7.7.49 (HIV Reverse Transcriptase)
RN  - EC 2.7.7.49 (RNA-Directed DNA Polymerase)
SB  - IM
MH  - Amino Acid Sequence
MH  - Base Sequence
MH  - DNA, Single-Stranded
MH  - Directed Molecular Evolution
MH  - Escherichia coli/*enzymology
MH  - HIV Reverse Transcriptase/genetics/metabolism
MH  - Ligands
MH  - Molecular Sequence Data
MH  - Nucleic Acid Conformation
MH  - Protein Binding
MH  - RNA/chemistry/*metabolism
MH  - RNA, Bacterial/chemistry/*metabolism
MH  - RNA-Directed DNA Polymerase/genetics/*metabolism
MH  - Recombinant Fusion Proteins/metabolism
MH  - Retroelements
MH  - Substrate Specificity
EDAT- 1999/10/26 00:00
MHDA- 1999/10/26 00:01
CRDT- 1999/10/26 00:00
PHST- 1999/10/26 00:00 [pubmed]
PHST- 1999/10/26 00:01 [medline]
PHST- 1999/10/26 00:00 [entrez]
AID - S0021-9258(19)51613-1 [pii]
AID - 10.1074/jbc.274.44.31236 [doi]
PST - ppublish
SO  - J Biol Chem. 1999 Oct 29;274(44):31236-44. doi: 10.1074/jbc.274.44.31236.