TY - JOUR
T1 - Two domains for splicing in the intron of the phage T4 thymidylate synthase (td) gene established by nondirected mutagenesis
AU - Hall, Dwight H.
AU - Povinelli, Christine M.
AU - Ehrenman, Karen
AU - Pedersen-Lane, Joan
AU - Chu, Frederick
AU - Belfort, Marlene
N1 - Funding Information:
D. H. H. acknowledges with deep appreciation Irwin Tessman and Salvador E. Luria in whose laboratories he isolated some of these mutants during his scientific training. We are most thankful to Dr. Chip Lawrence for aiding us in RNA secondary structure analysis of the mutants and for stimulating discussions. We thank Debbie Bell-Pedersen and R Scott Chandry for their helpfulness in the lab and for enthusiastic discussions. This work was supported by National Institutes of Health grant GM 33314 and National Science Foundation grants DMB-8502961 and DMB-8505527 to M. B. and NIH grants GM 16306, GM 24455, and GM 36714 to D. H. H.
PY - 1987/1/16
Y1 - 1987/1/16
N2 - Of 97 nondirected T4 thymidylate synthase-defective (td) mutations, 27 were mapped to the intron of the split td gene. Clustering of these intron mutations defined two domains that are functional in splicing, each within approximately 220 residues of the respective splice sites. Two selected mutations, tdN57 and tdN47, fell within phylogenetically conserved pairings, with tdN57 disrupting the exon I-internal guide pairing (P1) in the 5′ domain and tdN47 destabilizing the P9 helix in the 3′ domain. A splicing assay with synthetic oligonucleotides complementary to RNA junction sequences revealed processing defects for T4tdN57 and T4tdN47, both of which are impaired in cleavage at the 5′ and 3′ splice sites. Thus prokaryotic genetics facilitates association of specific residue changes with their consequences to splicing.
AB - Of 97 nondirected T4 thymidylate synthase-defective (td) mutations, 27 were mapped to the intron of the split td gene. Clustering of these intron mutations defined two domains that are functional in splicing, each within approximately 220 residues of the respective splice sites. Two selected mutations, tdN57 and tdN47, fell within phylogenetically conserved pairings, with tdN57 disrupting the exon I-internal guide pairing (P1) in the 5′ domain and tdN47 destabilizing the P9 helix in the 3′ domain. A splicing assay with synthetic oligonucleotides complementary to RNA junction sequences revealed processing defects for T4tdN57 and T4tdN47, both of which are impaired in cleavage at the 5′ and 3′ splice sites. Thus prokaryotic genetics facilitates association of specific residue changes with their consequences to splicing.
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U2 - 10.1016/0092-8674(87)90356-4
DO - 10.1016/0092-8674(87)90356-4
M3 - Article
C2 - 3791415
AN - SCOPUS:0023657389
VL - 48
SP - 63
EP - 71
JO - Cell
JF - Cell
SN - 0092-8674
IS - 1
ER -