Sequence-dependent conformational differences of small RNAs revealed by native gel electrophoresis

P J Beuning; M R Tessmer; C G Baumann; D A Kallick; K Musier-Forsyth

doi:10.1006/abio.1999.4223

Sequence-dependent conformational differences of small RNAs revealed by native gel electrophoresis

P J Beuning, M R Tessmer, C G Baumann, D A Kallick, K Musier-Forsyth

Biology

Research output: Contribution to journal › Article › peer-review

Abstract

In this study, we use native polyacrylamide gel electrophoresis and one-dimensional NMR spectroscopy to analyze small RNA hairpins containing a UUCG tetraloop. The aggregation state of one RNA 16-mer (5'-CGGCUUCGGUCGACCA-3') in the presence of Mg(2+) was confirmed by laser light scattering. Although it is widely known in the RNA field that some RNAs tend to aggregate, especially when present at high concentrations, the sequence elements responsible for this effect are rarely identified. In this work, we show that Mg(2+)-induced aggregation of the 16-mer RNA hairpin is sensitive to the presence of the 3'-terminal base and a specific 2'-hydroxyl group. Our study highlights the fact that even small changes in a particular RNA sequence can increase its tendency to undergo Mg(2+)-dependent aggregation in an unpredictable manner. Our analysis also shows that native gel electrophoresis is a sensitive probe of RNA conformation with the capability to detect differences apparently caused by subtle base stacking effects at the ends of helices.

Original language	English
Pages (from-to)	284-90
Number of pages	7
Journal	ANALYTICAL BIOCHEMISTRY
Volume	273
Issue number	2
DOIs	https://doi.org/10.1006/abio.1999.4223
Publication status	Published - 1999

Keywords

Base Sequence
Electrophoresis, Polyacrylamide Gel
Lasers
Magnetic Resonance Spectroscopy
Nucleic Acid Conformation
RNA
Scattering, Radiation

Access to Document

10.1006/abio.1999.4223

Cite this

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title = "Sequence-dependent conformational differences of small RNAs revealed by native gel electrophoresis",

abstract = "In this study, we use native polyacrylamide gel electrophoresis and one-dimensional NMR spectroscopy to analyze small RNA hairpins containing a UUCG tetraloop. The aggregation state of one RNA 16-mer (5'-CGGCUUCGGUCGACCA-3') in the presence of Mg(2+) was confirmed by laser light scattering. Although it is widely known in the RNA field that some RNAs tend to aggregate, especially when present at high concentrations, the sequence elements responsible for this effect are rarely identified. In this work, we show that Mg(2+)-induced aggregation of the 16-mer RNA hairpin is sensitive to the presence of the 3'-terminal base and a specific 2'-hydroxyl group. Our study highlights the fact that even small changes in a particular RNA sequence can increase its tendency to undergo Mg(2+)-dependent aggregation in an unpredictable manner. Our analysis also shows that native gel electrophoresis is a sensitive probe of RNA conformation with the capability to detect differences apparently caused by subtle base stacking effects at the ends of helices.",

keywords = "Base Sequence, Electrophoresis, Polyacrylamide Gel, Lasers, Magnetic Resonance Spectroscopy, Nucleic Acid Conformation, RNA, Scattering, Radiation",

author = "Beuning, {P J} and Tessmer, {M R} and Baumann, {C G} and Kallick, {D A} and K Musier-Forsyth",

year = "1999",

doi = "10.1006/abio.1999.4223",

language = "English",

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pages = "284--90",

journal = "ANALYTICAL BIOCHEMISTRY",

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T1 - Sequence-dependent conformational differences of small RNAs revealed by native gel electrophoresis

AU - Beuning, P J

AU - Tessmer, M R

AU - Baumann, C G

AU - Kallick, D A

AU - Musier-Forsyth, K

PY - 1999

Y1 - 1999

N2 - In this study, we use native polyacrylamide gel electrophoresis and one-dimensional NMR spectroscopy to analyze small RNA hairpins containing a UUCG tetraloop. The aggregation state of one RNA 16-mer (5'-CGGCUUCGGUCGACCA-3') in the presence of Mg(2+) was confirmed by laser light scattering. Although it is widely known in the RNA field that some RNAs tend to aggregate, especially when present at high concentrations, the sequence elements responsible for this effect are rarely identified. In this work, we show that Mg(2+)-induced aggregation of the 16-mer RNA hairpin is sensitive to the presence of the 3'-terminal base and a specific 2'-hydroxyl group. Our study highlights the fact that even small changes in a particular RNA sequence can increase its tendency to undergo Mg(2+)-dependent aggregation in an unpredictable manner. Our analysis also shows that native gel electrophoresis is a sensitive probe of RNA conformation with the capability to detect differences apparently caused by subtle base stacking effects at the ends of helices.

AB - In this study, we use native polyacrylamide gel electrophoresis and one-dimensional NMR spectroscopy to analyze small RNA hairpins containing a UUCG tetraloop. The aggregation state of one RNA 16-mer (5'-CGGCUUCGGUCGACCA-3') in the presence of Mg(2+) was confirmed by laser light scattering. Although it is widely known in the RNA field that some RNAs tend to aggregate, especially when present at high concentrations, the sequence elements responsible for this effect are rarely identified. In this work, we show that Mg(2+)-induced aggregation of the 16-mer RNA hairpin is sensitive to the presence of the 3'-terminal base and a specific 2'-hydroxyl group. Our study highlights the fact that even small changes in a particular RNA sequence can increase its tendency to undergo Mg(2+)-dependent aggregation in an unpredictable manner. Our analysis also shows that native gel electrophoresis is a sensitive probe of RNA conformation with the capability to detect differences apparently caused by subtle base stacking effects at the ends of helices.

KW - Base Sequence

KW - Electrophoresis, Polyacrylamide Gel

KW - Lasers

KW - Magnetic Resonance Spectroscopy

KW - Nucleic Acid Conformation

KW - RNA

KW - Scattering, Radiation

U2 - 10.1006/abio.1999.4223

DO - 10.1006/abio.1999.4223

M3 - Article

C2 - 10469499

SN - 0003-2697

VL - 273

SP - 284

EP - 290

JO - ANALYTICAL BIOCHEMISTRY

JF - ANALYTICAL BIOCHEMISTRY

IS - 2

ER -