Specific Recognition of Single-Stranded Regions in Ultraviolet-Irradiated and Heat-Denatured DNA by Tryptophan-Containing Peptides

Toulmé, Jean‐Jacques; Charlier, Michel; Hélène, Claude

doi:10.1073/pnas.71.8.3185

Cited by 76 publications

(31 citation statements)

References 14 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The observed preference of KWK for single-stranded polynucleotides (26,29) likely reflects the larger electrostatic contributions to the stability of these complexes, compared with doublestranded polynucleotides, and is consistent with our results. However, in those experiments, inner filter effects were evaluated from fluorescence quenching of the reactants in solutions with high salt concentrations where complexes were assumed to entirely dissociate.…”

Section: Discussionsupporting

confidence: 82%

“…Early fluorescence studies (26,29) reported that peptide KWK bound to single-and double-stranded polynucleotides at low salt concentrations primarily by electrostatic interactions. The observed preference of KWK for single-stranded polynucleotides (26,29) likely reflects the larger electrostatic contributions to the stability of these complexes, compared with doublestranded polynucleotides, and is consistent with our results.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Strandedness Discrimination in Peptide-Polynucleotide Complexes

Johnson

Mazarguil

Lopez

1996

Journal of Biological Chemistry

View full text Add to dashboard Cite

Preferential binding to single-or double-stranded nucleic acids is important for the activity of many proteins that process RNA and DNA. We have investigated the mechanism of strandedness discrimination with peptides derived from the putative DNA-binding domain of the RecA protein, a bacterial recombinase that modulates its affinity for single-stranded DNA by means of ATP binding and hydrolysis. Contributions of electrostatic and non-electrostatic interactions to binding of these peptides with polynucleotides were evaluated by fluorescence spectroscopy as a function of salt concentration and peptide charge. Binding of these peptides to single-and double-stranded nucleic acids was dominated by non-electrostatic interactions. Small electrostatic contributions selectively enhanced peptide complexation with single-stranded nucleic acids. Similar results were observed in control experiments carried out with tripeptides containing charged and aromatic amino acid residues. It was possible to modify the strandedness preference of peptide-polynucleotide complexes by changing electrostatic contributions to the binding free energy. These observations suggest a mechanism whereby some proteins that interact with DNA or RNA might determine and regulate their relative affinity for single-and double-stranded nucleic acids.Preferential binding to single-or double-stranded polynucleotides is characteristic of numerous proteins involved in DNA replication, transcription, DNA repair, and homologous recombination (1). Single-stranded binding proteins transiently stabilize single-stranded polynucleotides without NTP hydrolysis (2, 3). Helicases and recombinases may switch strandedness specificity by NTP-dependent mechanisms (4, 5). These proteins generally bind to nucleic acids cooperatively or as multimeric complexes. It is important to understand how these proteins recognize single-and double-stranded polynucleotides and how they change strandedness preference.Protein sequence motifs that recognize single-stranded DNA generally include both positively charged residues that can interact electrostatically with phosphate groups in the polynucleotide backbone and aromatic amino acids that can associate with nucleic acid bases via non-electrostatic interactions (2, 3, 6, 7). The L2 loop, a putative DNA-binding domain of the RecA protein, has charged and aromatic amino acid residues characteristic of single-stranded DNA-binding proteins; these general features are found in many recombinases (Fig. 1a) (8 -10). Genetic and biochemical experiments show that point mutations in this loop disrupt RecA protein function in vivo and in vitro (11)(12)(13)(14)(15) and modify the affinity of the RecA protein for single-stranded DNA (15). This loop is not resolved in the crystal structure of the RecA-ADP complex (16), which suggests that it may be flexible in the absence of DNA and that, at least to a certain extent, it might interact with DNA independently of the protein scaffolding. Hence, a peptide corresponding to the L2 loop may reproduce some o...

show abstract

Section: Discussionsupporting

confidence: 82%

Section: Discussionmentioning

confidence: 99%

Strandedness Discrimination in Peptide-Polynucleotide Complexes

Johnson

Mazarguil

Lopez

1996

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The interaction of tripeptide Lys-Trp-Lys with double-stranded and single-stranded DNA resulted in the quenching of the tryptophan fluorescence due to a stacking of the indole ring with bases [54]. A comparison of the association constants, K , showed that the affinity of TP4 for double-stranded DNA was about 1.8-fold higher than that for single-stranded DNA.…”

Section: Tp4 Is Composed Of 138 Amino Acid Residues Its Calculated Mmentioning

confidence: 99%

The Amino Acid Sequence and Interaction with the Nucleosome Core DNA of Transition Protein 4 from Boar Late Spermatid Nuclei

Akama¹,

Ichimura²,

Sato³

et al. 1995

European Journal of Biochemistry

View full text Add to dashboard Cite

The primary structure of transition protein 4 (TP4) from boar late spermatid nuclei was determined by automated Edman degradation of S-pyridylethylated protein and of peptides generated by cleavage with Staphylococcus aureus V8 protease, lysyl endopeptidase and CNBr. Boar TP4 is a basic protein consisting of a highly basic amino-terminal half (residues 1-73) and a less basic carboxy-terminal half (residues 74-1 38). The latter half includes a highly hydrophobic segment, a four-times tandemly repeated sequence, N(G)QNKR(K)X, and a carboxy-terminal segment containing Trpl26. Ultraviolet absorption and CD spectra of TP4 -rat-liver-nucleosome-core-DNA (double-stranded DNA) complexes suggest a TP4-induced local melting of DNA. Although at 1 mM NaCl TP4 brought about a slight stabilization of the DNA against thermal melting, a destabilization of the DNA was observed at 50 mM NaCl. From the results of quenching of tryptophan (Trp126) fluorescence of TP4 upon its binding to double-stranded and single-stranded boar liver nucleosome-core DNA at 50 mM NaCl, the apparent association constants for the binding of TP4 to double-stranded and single-stranded DNA were calculated to be 7.3X10' M-' and 4.1 X 10' M -', respectively. These results suggest that TP4, having different domain structures from TP1-3 and a higher affinity for double-stranded DNA, induces a local destabilization of DNA probably through the stacking of Trp126 with nucleic acid bases.Keywords: transition protein ; amino acid sequence; DNA-protein interaction; late spermatid nuclei.The mechanism of transformation of nucleosome-type chromatin into a nucleoprotamine fiber is rather poorly understood. A direct transition from a nucleosome-type chromatin organization into a nucleoprotamine fiber operates in most species with the exception of mammals [I, 21. In mammals, nucleosomal histones are transiently replaced by small basic proteins called transition proteins (TPI -4) and, finally, by protamines [3-61. At the same time, transformation of the nucleosomal-type chromatin into a smooth chromatin fiber, initiation of chromatin condensation and cessation of transcription occur [7-121. The amino acid sequences of TPI and TP2, and the nucleotide sequences of their genes are known [ 13 -161, and attempts have been made to understand how they are transcriptionally and translationally regulated [17, 181. It has been postulated that rat TP1 is a DNA melting protein, mediated through the intercalation of its tyrosine residue between the nucleic acid bases [19], that it destabilizes the compact nucleosome core particles 1201, and that rat TP2, with two possible zinc fingers in the amino- terminal region, is a DNA stabilizing protein [21, 221. Recently, we have developed methods for isolating intact boar TP1, TP3 and TP4 123, 241, and have reported that boar TP3 has 27% sequence similarity to boar TPI [25]. In this paper, we describe the amino acid sequence of boar TP4 and the nature of interaction of TP4 with nucleosome-core DNA in vitro. MATERIALS AND METHODSMaterials. sta...

show abstract

“…For the tripeptide lysyltryptophyl-a-lysine (Lys-Trp-Lys) we demonstrated that stacking interactions were favored in complexes with single-stranded polynucleotides as compared with double-stranded DNA (12,13). Moreover, this peptide was able to bind more efficiently to locally destabilized regions of DNA after modification by UV irradiation or by N-acetoxy-N-2-acetylaminofluorene (13)(14)(15).…”

mentioning

confidence: 99%

Specific recognition of apurinic sites in DNA by a tryptophan-containing peptide.

Behmoaras¹,

Toulmé²,

Hélène³

1981

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

We have used fluorescence spectroscopy to study the binding of lysyltryptophyl-a-lysine (Lys-Trp-Lys) to DNA modified by dimethyl sulfate before and after depurination and strand breakage. Quenching of tryptophan fluorescence increased upon association of the peptide with modified DNA as compared with native DNA. We have demonstrated that this quenching is related to a preferential stacking of the indole ring with nucleic acid bases in damaged regions. Stacking increased in the following order: methylated DNA < DNA with strand breaks at apurinic sites << apurinic DNA. For apurinic DNA, the overall association constant of Lys-Trp-Lys was increased by more than two orders of magnitude as compared to native DNA. Enhancement of the affinity of the tripeptide for an apurinic site requires the integrity of the phosphodiester bond. Single-strand cleavage at an apurinic site leads to a marked decrease of the association constant. The peptide Lys-Trp-Lys is therefore able to recognize destabilized regions in the vicinity of a lesion and to discriminate between different confignrations of the damaged region. These results are discussed with respect to the role that stacking interactions could play in the specificity of recognition of DNA alterations by enzymes involved in DNA repair mechanisms.Alkylating agents produce several types of alterations in nucleic acids, especially in the base moeities of the polynucleotide chain (1). N7-Methylguanine and N3-methyladenine are the two major lesions (76% and 18%, respectively) in DNA treated with dimethyl sulfate (2). The glycosidic bond of these methylated bases is labilized as compared to unmodified bases, leading to the formation of apurinic sites. Depurination of DNA occurs spontaneously at a significant rate even under physiological conditions (3). In vivo these lesions (i.e., methylated bases, except N7-methylguanine, or apurinic sites) have to be repaired by the cell to preserve the integrity of its genome. This is done by specific enzymes: e.g., glycosylases, apurinic endonucleases, and insertases (4-9). Nothing is known about the mechanism of recognition of DNA defects by these enzymes.We have shown previously that oligopeptides containing aromatic amino acids were able to form stacked complexes with nucleic acids (10, 11). For the tripeptide lysyltryptophyl-a-lysine (Lys-Trp-Lys) we demonstrated that stacking interactions were favored in complexes with single-stranded polynucleotides as compared with double-stranded DNA (12, 13). Moreover, this peptide was able to bind more efficiently to locally destabilized regions of DNA after modification by UV irradiation or by N-acetoxy-N-2-acetylaminofluorene (13-15).We have studied the binding of Lys-Trp-Lys to methylated DNA (MeDNA) and to DNA containing apurinic sites (ap-DNA) in order to determine whether this tripeptide was able to discriminate between native and chemically modified DNA. We have shown that removal of purines introduces strong binding sites as a result of efficient stacking of the tryptophyl residue....

show abstract

Specific Recognition of Single-Stranded Regions in Ultraviolet-Irradiated and Heat-Denatured DNA by Tryptophan-Containing Peptides

Cited by 76 publications

References 14 publications

Strandedness Discrimination in Peptide-Polynucleotide Complexes

Strandedness Discrimination in Peptide-Polynucleotide Complexes

The Amino Acid Sequence and Interaction with the Nucleosome Core DNA of Transition Protein 4 from Boar Late Spermatid Nuclei

Specific recognition of apurinic sites in DNA by a tryptophan-containing peptide.

Contact Info

Product

Resources

About