DNA hairpin loops in solution. Correlation between primary structure, thermostability and reactivity with single-strand-specific nuclease from mung bean

Xodo, Luigi E.; Manzini, Giorgio; Quadrifoglio, Franco; Marel, Gijs van der; Boom, Jacques van

doi:10.1093/nar/19.7.1505

Cited by 58 publications

(36 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…This prediction has been verified by previous studies showing that at least two single-strand-specific endonucleases, SI nuclease and mung bean nuclease (MBN), are capable of introducing nicks in the vicinity ofhairpin loops (23)(24)(25). However, all but one of the hairpins examined in those studies contained terminal loops with 3-8 non-complementary nucleotides.…”

Section: Introductionsupporting

confidence: 70%

Hairpin opening by single-strand-specific nucleases

Kabotyanski

Zhu²,

Kallick

et al. 1995

Nucl Acids Res

View full text Add to dashboard Cite

DNA molecules with covalently sealed (hairpin) ends are probable intermediates in V(D)J recombination. According to current models hairpin ends are opened to produce short single-stranded extensions that are thought to be precursors of a particular type of extra nucleotides, termed P nucleotides, which are frequently present at recombination junctions. Nothing is known about the activites responsible for hairpin opening. We have used two single-strand-specIfIc nucleases to explore the effects of loop sequence on the hairpin opening reaction. Here we show that a variety of hairpin ends are opened by P1 nuclease and mung bean nuclease (MBN) to leave short, 1-2 nt single-stranded extensions. Analysis of 22 dIfferent hairpin sequences demonstrates that the terminal 4 nt of the hairpin loop strongly influence the sites of cleavage. Correlation of the nuclease digestion patterns with structura (NMR) data for some of the hairpin loops studied here provides new insights into the structural features recognized by these enzymes.

show abstract

Section: Introductionsupporting

confidence: 70%

Hairpin opening by single-strand-specific nucleases

Kabotyanski

Zhu²,

Kallick

et al. 1995

Nucl Acids Res

View full text Add to dashboard Cite

show abstract

“…A DNA hairpin is made up of two parts: a base-paired stem and a single-stranded loop connecting the two halves of the stem. Structural studies have shown that the loop of fully complementary hairpins generally adopts a 2-or 4-unpaired-nucleotide structure, due to the tremendous steric constraints imposed by the bonding of the last two nucleotides (29,35,36). A DNA hairpin has therefore to be seen as a dynamic equilibrium adopting the more favorable energy conformation under a given steric stress.…”

Section: Discussionmentioning

confidence: 99%

“…Numerous factors are involved in the counterbalance of stabilities of the loop and the stem regions and determine the predominant conformations of the hairpin (5,10). One very relevant factor is the effect of the base sequence of DNA hairpins on hairpin stability and loop folding (2,13,29,36). For example, Blommers et al have shown that the same hairpin will have a loop of 2 or 4 nucleotides upon a single nucleotide change in position N 1 (2).…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, several studies have shown that the secondary structures of DNA hairpins can affect enzyme recognition and cleavage. For example, it has been shown that the activity and sites of cleavage of single strandspecific nucleases are dependent on the structure and size of the hairpin loop and on structural distortions of nucleotides in the vicinity (14,36). Also, Froelich-Ammon et al (8) have shown that topoisomerase II, which is believed to mediate nonhomologous recombination events in the cell, cleaves DNA hairpins in a site-specific fashion.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Nucleotide Deletion and P Addition in V(D)J Recombination: a Determinant Role of the Coding-End Sequence†

Nadel

Feeney

1997

Molecular and Cellular Biology

View full text Add to dashboard Cite

During V(D)J recombination, the coding ends to be joined are extensively modified. Those modifications, termed coding-end processing, consist of removal and addition of various numbers of nucleotides. We previously showed in vivo that coding-end processing is specific for each coding end, suggesting that specific motifs in a coding-end sequence influence nucleotide deletion and P-region formation. In this study, we created a panel of recombination substrates containing actual immunoglobulin and T-cell receptor coding-end sequences and dissected the role of each motif by comparing its processing pattern with those of variants containing minimal nucleotide changes from the original sequence. Our results demonstrate the determinant role of specific sequence motifs on coding-end processing and also the importance of the context in which they are found. We show that minimal nucleotide changes in key positions of a coding-end sequence can result in dramatic changes in the processing pattern. We propose that each coding-end sequence dictates a unique hairpin structure, the result of a particular energy conformation between nucleotides organizing the loop and the stem, and that the interplay between this structure and specific sequence motifs influences the frequency and location of nicks which open the coding-end hairpin. These findings indicate that the sequences of the coding ends determine their own processing and have a profound impact on the development of the primary B-and T-cell repertoires.The large repertoire of immunoglobulin (Ig) and T-cell receptor (TCR) variable regions is generated during V(D)J recombination by two main sources of diversity (34). Combinatorial diversity derives from the assembly of one of the many variable (V), diversity (D), and joining (J) gene segments to form a V(D)J exon and, subsequently, from the pairing of the two chains of the heterodimeric receptor. Junctional diversity is created by extensive nucleotide modification at the junctions during the assembly of the gene segments. V(D)J recombination is therefore the earliest mechanism generating diversity in the primary B-and T-cell repertoires.The proper rearrangement of the various gene segments requires the presence of recombination signal sequences (RSS) adjacent to each coding segment. This signal consists of conserved heptamer and nonamer sequences separated by a nonconserved spacer region of 12 or 23 bp (called 12-bp RSS and 23-bp RSS, respectively) (1, 12). The initial DNA break is performed by the RAG1-RAG2 complex, which binds to the RSS and makes a single-strand cut at the 5Ј end of the heptamer at the precise border of the coding end and the RSS (23). The phosphodiester bond of the complementary strand opposite to the initial nick is then attacked by direct transesterification, resulting in a covalently sealed hairpin coding end and a blunt signal end.To proceed through the recombination process, the DNA hairpins formed by the coding ends have to be resolved into free ends, which is presumably done by double-or singlestra...

show abstract

“…Consequently, one, two, three, and four residue(s) of analogue X were planned to be incorporated into the loop moiety. The hexameric stem hairpin structure would be suitable for this study, because it is known to form stable hairpins with a loop consisting of two, three, or four thymidine residues (28). Thus, we designed the hairpinforming ODNs 5′-d[CGAACG-X n -CGTTCG]-3′ (I-1, n ) 1; I-2, n ) 2; I-3, n ) 3; I-4, n ) 4), shown in Figure 3, for this study.…”

Section: Resultsmentioning

confidence: 99%

Oligodeoxynucleotides Having a Loop Consisting of 3‘-Deoxy-4‘-C-(2-hydroxyethyl)thymidines Form Stable Hairpins

Yamamoto¹,

Shuto²,

Tsutsumi³

et al. 2004

Biochemistry

View full text Add to dashboard Cite

Components that form stable hairpin loops are highly useful for the development of functional DNA and RNA molecules. We have designed and synthesized a sugar-modified thymidine analogue, 3'-deoxy-4'-C-(2-hydroxyethyl)thymidine (X), as a nucleosidic loop component stabilizing the hairpin structure. The ODNs I-1-4, 5'-d[CGAACG-X(n)-CGTTCG]-3' (I-1, n = 1; I-2, n = 2; I-3, n = 3; I-4, n = 4), forming the hairpin loop structures, of which the loop moiety consisted of the analogue X, and also the corresponding unmodified ODNs II-1-4, 5'-d[CGAACG-T(n)-CGTTCG]-3' (II-1, n = 1; II-2, n = 2;II-3, n = 3; II-4, n = 4), having a thymidine loop, were synthesized by the phosphoramidite method. The melting temperatures (T(m)) of the ODNs I-1-4 containing X in the loop moiety at 5 microM were 67.1, 68.1, 73.0, and 69.3 degrees C, respectively, and those of the control natural ODNs II-1-4 were 65.3, 67.0, 69.2, and 68.8 degrees C, respectively. Thus, the ODNs I-1-4 formed a more thermally stable hairpin than the corresponding unmodified ODNs II-1-4 having an equal number of loop residues. The hairpin structures of the modified ODNs I-1-4 and the unmodified ODNs II-1-4 were investigated by CD spectroscopy and molecular mechanics calculations. These results showed that the 4'-branched nucleoside X can stabilize hairpin structures when it is present in the loop moiety, probably due to the flexibility of the one-carbon-elongated 4'-branched structure.

show abstract

DNA hairpin loops in solution. Correlation between primary structure, thermostability and reactivity with single-strand-specific nuclease from mung bean

Cited by 58 publications

References 33 publications

Hairpin opening by single-strand-specific nucleases

Hairpin opening by single-strand-specific nucleases

Nucleotide Deletion and P Addition in V(D)J Recombination: a Determinant Role of the Coding-End Sequence†

Oligodeoxynucleotides Having a Loop Consisting of 3‘-Deoxy-4‘-C-(2-hydroxyethyl)thymidines Form Stable Hairpins

Contact Info

Product

Resources

About