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Titolo:
Structural basis of the enhanced stability of a mutant ribozyme domain anda detailed view of RNA-solvent interactions
Autore:
Juneau, K; Podell, E; Harrington, DJ; Cech, TR;
Indirizzi:
Univ Colorado, Dept Chem & Biochem, Boulder, CO 80309 USA Univ Colorado Boulder CO USA 80309 Chem & Biochem, Boulder, CO 80309 USA Univ Colorado, Howard Hughes Med Inst, Boulder, CO 80309 USA Univ Colorado Boulder CO USA 80309 Hughes Med Inst, Boulder, CO 80309 USA
Titolo Testata:
STRUCTURE
fascicolo: 3, volume: 9, anno: 2001,
pagine: 221 - 231
SICI:
0969-2126(20010307)9:3<221:SBOTES>2.0.ZU;2-V
Fonte:
ISI
Lingua:
ENG
Soggetto:
PHENYLALANINE TRANSFER-RNA; GROUP-I RIBOZYME; DELTA VIRUS RIBOZYME; CRYSTAL-STRUCTURE; TERTIARY STRUCTURE; ANGSTROM RESOLUTION; HAMMERHEAD RIBOZYME; HYDROGEN-BONDS; BINDING DOMAIN; PROTEIN;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
52
Recensione:
Indirizzi per estratti:
Indirizzo: Cech, TR Univ Colorado, Dept Chem & Biochem, Boulder, CO 80309 USA Univ Colorado Boulder CO USA 80309 iochem, Boulder, CO 80309 USA
Citazione:
K. Juneau et al., "Structural basis of the enhanced stability of a mutant ribozyme domain anda detailed view of RNA-solvent interactions", STRUCTURE, 9(3), 2001, pp. 221-231

Abstract

Background: The structure of P4-P6, a 160 nucleotide domain of the self-splicing Tetrahymena thermophila intron, was solved previously. Mutants of the P4-P6 RNA that form a more stable tertiary structure in solution were recently isolated by successive rounds of in vitro selection and amplification. Results: We show that a single-site mutant (Delta C209) possessing greatertertiary stability than wild-type P4-P6 also crystallizes much more rapidly and under a wider variety of conditions. The crystal structure provides asatisfying explanation for the increased stability of the mutant; the deletion of C209 allows the adjacent bulged adenine to enter the P4 helix and form an A-G base pair, presumably attenuating the conformational flexibilityof the helix. The structure of another mutant (Delta A210) was also solvedand supports this interpretation. The crystals of Delta C209 diffract to ahigher resolution limit than those of wild-type RNA (2.25 Angstrom versus 2.8 Angstrom), allowing assignment of innersphere and outersphere coordination contacts for 27 magnesium ions. Structural analysis reveals an intricate solvent scaffold with a preponderance of ordered water molecules on the inside rather than the surface of the folded RNA domain. Conclusions: In vitro evolution facilitated the identification of a highlystable, structurally homogeneous mutant RNA that was readily crystallizable. Analysis of the structure suggests that improving RNA secondary structure can stabilize tertiary structure and perhaps promote crystallization. In addition, the higher resolution model provides new details of metal ion-RNAinteractions and identifies a core of ordered water molecules that may be integral to RNA tertiary structure formation.

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Documento generato il 03/07/20 alle ore 01:40:12