噬菌体R17 RNA中尿苷的光水合作用。I. 尿苷光水合作用的还原测定法。

Photohydration of uridine in the RNA of coliphage R17. I. Reductive assay for uridine photohydration.

作者信息

Cerutti P A, Miller N, Pleiss M G, Remsen J F, Ramsay W J

出版信息

Proc Natl Acad Sci U S A. 1969 Oct;64(2):731-8. doi: 10.1073/pnas.64.2.731.

DOI:10.1073/pnas.64.2.731

PMID:5261044

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC223405/

Abstract

Photohydration of uridine in the RNA of E. coli bacteriophage R17 has been investigated with the help of a new chemical method which avoids digestion of the irradiated polymer. The uridine photohydrates are reductively cleaved with sodium borotritiide and the radioactively labeled fragment, 1,3-propane-diol-(3)H(alpha), (3)H(alpha), (3)H(gamma), formed in this reaction is isolated and used as a measure for the extent of uridine photohydration in the irradiated RNA. The influence of the conformational state of R17-RNA on the cross section of uridine photohydration (sigma(H)) was investigated. The cross section for irradiation at 280 mmu at 25 degrees was largest in 6 M urea (sigma(H) = 0.053), slightly smaller in low salt (<10(-3)M Na(+); sigma(H) = 0.050), and substantially smaller in 0.15 M Na(+) (sigma(H) = 0.038). The closeness of the values for sigma(H) in urea and low salt indicates that a majority of the uridine residues in R17-RNA in the low salt medium at 25 degrees are not hydrogen bonded and destacked. The significant suppression of photohydration in 0.15 M Na(+) on the other hand shows that a substantial portion of the uridine residues in R17-RNA are participating in base pairing and base stacking under these conditions. Our results demonstrate that studies of uridine photohydration may, beyond their importance to photobiology, yield information specifically about the conformational state of the uridine-rich regions of a RNA, information which cannot be readily obtained by other techniques.

摘要

借助一种新的化学方法，对大肠杆菌噬菌体R17 RNA中的尿苷光水合作用进行了研究，该方法可避免对辐照后的聚合物进行消化。用硼氢化三氚钠对尿苷光水合物进行还原裂解，并分离出此反应中形成的放射性标记片段1,3 - 丙二醇 -（3）H（α），（3）H（α），（3）H（γ），将其用作衡量辐照RNA中尿苷光水合程度的指标。研究了R17 - RNA的构象状态对尿苷光水合截面（σ(H)）的影响。在25℃下，280毫微米处的辐照截面在6 M尿素中最大（σ(H)=0.053），在低盐（<10⁻³ M Na⁺；σ(H)=0.050）时稍小，而在0.15 M Na⁺中则小得多（σ(H)=0.038）。尿素和低盐条件下σ(H)值相近，表明在25℃的低盐介质中，R17 - RNA中的大多数尿苷残基未形成氢键且未堆积。另一方面，0.15 M Na⁺中光水合作用的显著抑制表明，在这些条件下，R17 - RNA中的相当一部分尿苷残基参与了碱基配对和碱基堆积。我们的结果表明，尿苷光水合作用的研究除了对光生物学具有重要意义外，还可能产生有关RNA富含尿苷区域构象状态的特定信息，而这些信息是其他技术难以轻易获得的。