Secondary structure and stability of the selenocysteine insertion sequences (SECIS) for human thioredoxin reductase and glutathione peroxidase.
We have used high resolution NMR and thermodynamics to characterize the secondary structure and stability of the selenocysteine insertion sequences (SECIS) of human glutathione peroxidase (58 nt) and thioredoxin reductase (51 nt). These sequences are members of the two classes of SECIS recently identified with two distinct structures capable of directing selenocysteine incorporation into proteins in eukaryotes. UV melting experiments showed a single cooperative and reversible transition for each RNA, which indicates the presence of stable secondary structures. Despite their large size, the RNAs gave well resolved NMR spectra for the exchangeable protons. Using NOESY, the imino protons as well as the cytosine amino protons of all of the Watson-Crick base pairs were assigned. In addition, a number of non-canonical base pairs including the wobble G.U pairs were identified. The interbase-pair NOEs allowed definition of the hydrogen-bonded structure of the oligonucleotides, providing an experimental model of the secondary structure of these elements. The derived secondary structures are consistent with several features of the predicted models, but with some important differences, especially regarding the conserved sequence motifs.
Ramos
Andres
A
Division of Molecular Structure, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, UK.
Lane
Andrew N
AN
Hollingworth
David
D
Fan
Teresa W-M
TW
D001483:Base Sequence; D005979:Glutathione Peroxidase / Q000235:genetics*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D020871:RNA Stability; D038621:Regulatory Sequences, Ribonucleic Acid; D017279:Selenocysteine / Q000378:metabolism*; D013880:Thioredoxin-Disulfide Reductase / Q000235:genetics*
D038621:Regulatory Sequences, Ribonucleic Acid; D017279:Selenocysteine; D012313:RNA; D005979:Glutathione Peroxidase; D013880:Thioredoxin-Disulfide Reductase