MersV1, Istex, Corpus, bibRecord, 001D57

Methylphosphonate mapping of phosphate contacts critical for RNA recognition by the human immunodeficiency virus tat and rev proteins

Identifieur interne : 001D57 ( Istex/Corpus ); précédent : 001D56; suivant : 001D58

Methylphosphonate mapping of phosphate contacts critical for RNA recognition by the human immunodeficiency virus tat and rev proteins

Auteurs : Clare E. Pritchard ; Jane A. Grasby ; François Hamy ; Anthony M. Zacharek ; Mohinder Singh ; Jonathan Karn ; Michael J. Gait

Source :

Nucleic Acids Research [ 0305-1048 ] ; 1994.

RBID : ISTEX:C14DEE447A84C35F30C19F42ECFED28A3773A037

Abstract

The HIV-1 regulatory proteins tat and rev are both RNA binding proteins which recognize sequences in duplex RNA which are close to structural distortions. Here we identify phosphate contacts which are critical for each binding reaction by use of a new method. Model RNA binding sites are constructed carrying substitutions of individual phosphodiesters by uncharged methylphosphonate derivatives isolated separately as ftp and Sp diastereoisomers and tested for protein binding by competition assays. In the binding of tat to the trans-activation response region (TAR), three phosphates, P21 and P22 which are adjacent to the U-rich bulge and P40 on the opposite strand, are essential and in each case both isomers inhibit binding. Similarly, in the interaction between the HIV-1 rev protein and the revresponsive element (RRE) both methylphosphonate isomers at P103, P104, P124 and P125 interfere with rev binding. At P106, only the Rp methylphosphonate isomer is impaired in rev binding ability and it is proposed that the Rp oxygen is hydrogen-bonded to an uncharged amino acid or to a main chain hydrogen atom. Synthetic chemistry techniques also provide evidence for the conformations of non-Watson - Crick G106:G129 and G105:A131 base-pairs in the RRE ‘bubble’ structure upon rev binding. Almost all functional groups on the 5 bulged residues in the bubble have been ruled out as sites of contact with rev but, by contrast, the N7-positions of each G residue in the flanking base-pairs are identified as sites of likely hydrogen-bonding to rev. The results show that both tat and rev recognize the major groove of distorted RNA helixes and that both proteins make specific contacts with phosphates which are displaced from the sites of base-pair contact.

Url:

https://api.istex.fr/ark:/67375/HXZ-SCD9CQ9K-G/fulltext.pdf

DOI: 10.1093/nar/22.13.2592

Links to Exploration step

ISTEX:C14DEE447A84C35F30C19F42ECFED28A3773A037

Le document en format XML

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<front><div type="abstract">The HIV-1 regulatory proteins tat and rev are both RNA binding proteins which recognize sequences in duplex RNA which are close to structural distortions. Here we identify phosphate contacts which are critical for each binding reaction by use of a new method. Model RNA binding sites are constructed carrying substitutions of individual phosphodiesters by uncharged methylphosphonate derivatives isolated separately as ftp and Sp diastereoisomers and tested for protein binding by competition assays. In the binding of tat to the trans-activation response region (TAR), three phosphates, P21 and P22 which are adjacent to the U-rich bulge and P40 on the opposite strand, are essential and in each case both isomers inhibit binding. Similarly, in the interaction between the HIV-1 rev protein and the revresponsive element (RRE) both methylphosphonate isomers at P103, P104, P124 and P125 interfere with rev binding. At P106, only the Rp methylphosphonate isomer is impaired in rev binding ability and it is proposed that the Rp oxygen is hydrogen-bonded to an uncharged amino acid or to a main chain hydrogen atom. Synthetic chemistry techniques also provide evidence for the conformations of non-Watson - Crick G106:G129 and G105:A131 base-pairs in the RRE ‘bubble’ structure upon rev binding. Almost all functional groups on the 5 bulged residues in the bubble have been ruled out as sites of contact with rev but, by contrast, the N7-positions of each G residue in the flanking base-pairs are identified as sites of likely hydrogen-bonding to rev. The results show that both tat and rev recognize the major groove of distorted RNA helixes and that both proteins make specific contacts with phosphates which are displaced from the sites of base-pair contact.</div>
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<abstract>The HIV-1 regulatory proteins tat and rev are both RNA binding proteins which recognize sequences in duplex RNA which are close to structural distortions. Here we identify phosphate contacts which are critical for each binding reaction by use of a new method. Model RNA binding sites are constructed carrying substitutions of individual phosphodiesters by uncharged methylphosphonate derivatives isolated separately as ftp and Sp diastereoisomers and tested for protein binding by competition assays. In the binding of tat to the trans-activation response region (TAR), three phosphates, P21 and P22 which are adjacent to the U-rich bulge and P40 on the opposite strand, are essential and in each case both isomers inhibit binding. Similarly, in the interaction between the HIV-1 rev protein and the revresponsive element (RRE) both methylphosphonate isomers at P103, P104, P124 and P125 interfere with rev binding. At P106, only the Rp methylphosphonate isomer is impaired in rev binding ability and it is proposed that the Rp oxygen is hydrogen-bonded to an uncharged amino acid or to a main chain hydrogen atom. Synthetic chemistry techniques also provide evidence for the conformations of non-Watson - Crick G106:G129 and G105:A131 base-pairs in the RRE ‘bubble’ structure upon rev binding. Almost all functional groups on the 5 bulged residues in the bubble have been ruled out as sites of contact with rev but, by contrast, the N7-positions of each G residue in the flanking base-pairs are identified as sites of likely hydrogen-bonding to rev. The results show that both tat and rev recognize the major groove of distorted RNA helixes and that both proteins make specific contacts with phosphates which are displaced from the sites of base-pair contact.</abstract>
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<profileDesc><abstract><p>The HIV-1 regulatory proteins tat and rev are both RNA binding proteins which recognize sequences in duplex RNA which are close to structural distortions. Here we identify phosphate contacts which are critical for each binding reaction by use of a new method. Model RNA binding sites are constructed carrying substitutions of individual phosphodiesters by uncharged methylphosphonate derivatives isolated separately as ftp and Sp diastereoisomers and tested for protein binding by competition assays. In the binding of tat to the <hi rend="italic">trans</hi>
-activation response region (TAR), three phosphates, P21 and P22 which are adjacent to the U-rich bulge and P40 on the opposite strand, are essential and in each case both isomers inhibit binding. Similarly, in the interaction between the HIV-1 rev protein and the revresponsive element (RRE) both methylphosphonate isomers at P103, P104, P124 and P125 interfere with rev binding. At P106, only the <hi rend="italic">R</hi>
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<article-title>Methylphosphonate mapping of phosphate contacts critical for RNA recognition by the human immunodeficiency virus tat and rev proteins</article-title>

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<contrib contrib-type="author">

<name><surname>Pritchard</surname>
<given-names>Clare E.</given-names>
</name>
<xref ref-type="fn" rid="fn1"><sup>+</sup>
</xref>

</contrib>

<contrib contrib-type="author">

<name><surname>Grasby</surname>
<given-names>Jane A.</given-names>
</name>

</contrib>

<contrib contrib-type="author">

<name><surname>Hamy</surname>
<given-names>François</given-names>
</name>
<xref ref-type="fn" rid="fn2"><sup>§</sup>
</xref>

</contrib>

<contrib contrib-type="author">

<name><surname>Zacharek</surname>
<given-names>Anthony M.</given-names>
</name>
<xref ref-type="aff" rid="au1"><sup>¶</sup>
</xref>

</contrib>

<contrib contrib-type="author">

<name><surname>Singh</surname>
<given-names>Mohinder</given-names>
</name>

</contrib>

<contrib contrib-type="author">

<name><surname>Karn</surname>
<given-names>Jonathan</given-names>
</name>

</contrib>

<contrib contrib-type="author">

<name><surname>Gait</surname>
<given-names>Michael J.</given-names>
</name>
<xref ref-type="corresp" rid="cor1"><sup>*</sup>
</xref>

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<aff><institution>MRC Laboratory of Molecular Biology</institution>
 <addr-line>Hills Road, Cambridge CB2 2QH, UK</addr-line>
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<aff id="au1"><sup>¶</sup>
<institution>Department of Biochemical Sciences, Harvard University</institution>
 <addr-line>Cambridge, MA, USA</addr-line>
</aff>

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<corresp id="cor1"><sup>*</sup>
To whom correspondence should be addressed</corresp>

<fn id="fn1"><p><sup>+</sup>
Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK</p>
</fn>

<fn id="fn2"><p><sup>§</sup>
Ciba-Geigy AG, CH-4002 Basel, Switzerland</p>
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<day>11</day>

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<year>1994</year>

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<year>1994</year>

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<year>1994</year>

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<copyright-statement>© 1994 Oxford University Press</copyright-statement>

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<abstract>

<p>The HIV-1 regulatory proteins tat and rev are both RNA binding proteins which recognize sequences in duplex RNA which are close to structural distortions. Here we identify phosphate contacts which are critical for each binding reaction by use of a new method. Model RNA binding sites are constructed carrying substitutions of individual phosphodiesters by uncharged methylphosphonate derivatives isolated separately as ftp and Sp diastereoisomers and tested for protein binding by competition assays. In the binding of tat to the <italic>trans</italic>
-activation response region (TAR), three phosphates, P21 and P22 which are adjacent to the U-rich bulge and P40 on the opposite strand, are essential and in each case both isomers inhibit binding. Similarly, in the interaction between the HIV-1 rev protein and the revresponsive element (RRE) both methylphosphonate isomers at P103, P104, P124 and P125 interfere with rev binding. At P106, only the <italic>R</italic>
p methylphosphonate isomer is impaired in rev binding ability and it is proposed that the <italic>R</italic>
p oxygen is hydrogen-bonded to an uncharged amino acid or to a main chain hydrogen atom. Synthetic chemistry techniques also provide evidence for the conformations of non-Watson - Crick G<sub>106</sub>
:G<sub>129</sub>
 and G<sub>105</sub>
:A<sub>131</sub>
 base-pairs in the RRE ‘bubble’ structure upon rev binding. Almost all functional groups on the 5 bulged residues in the bubble have been ruled out as sites of contact with rev but, by contrast, the <italic>N</italic>
<sup>7</sup>
-positions of each G residue in the flanking base-pairs are identified as sites of likely hydrogen-bonding to rev. The results show that both tat and rev recognize the major groove of distorted RNA helixes and that both proteins make specific contacts with phosphates which are displaced from the sites of base-pair contact.</p>

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<abstract>The HIV-1 regulatory proteins tat and rev are both RNA binding proteins which recognize sequences in duplex RNA which are close to structural distortions. Here we identify phosphate contacts which are critical for each binding reaction by use of a new method. Model RNA binding sites are constructed carrying substitutions of individual phosphodiesters by uncharged methylphosphonate derivatives isolated separately as ftp and Sp diastereoisomers and tested for protein binding by competition assays. In the binding of tat to the trans-activation response region (TAR), three phosphates, P21 and P22 which are adjacent to the U-rich bulge and P40 on the opposite strand, are essential and in each case both isomers inhibit binding. Similarly, in the interaction between the HIV-1 rev protein and the revresponsive element (RRE) both methylphosphonate isomers at P103, P104, P124 and P125 interfere with rev binding. At P106, only the Rp methylphosphonate isomer is impaired in rev binding ability and it is proposed that the Rp oxygen is hydrogen-bonded to an uncharged amino acid or to a main chain hydrogen atom. Synthetic chemistry techniques also provide evidence for the conformations of non-Watson - Crick G106:G129 and G105:A131 base-pairs in the RRE ‘bubble’ structure upon rev binding. Almost all functional groups on the 5 bulged residues in the bubble have been ruled out as sites of contact with rev but, by contrast, the N7-positions of each G residue in the flanking base-pairs are identified as sites of likely hydrogen-bonding to rev. The results show that both tat and rev recognize the major groove of distorted RNA helixes and that both proteins make specific contacts with phosphates which are displaced from the sites of base-pair contact.</abstract>
<note type="footnotes">+Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK</note>
<note type="author-notes">*To whom correspondence should be addressed</note>
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Methylphosphonate mapping of phosphate contacts critical for RNA recognition by the human immunodeficiency virus tat and rev proteins

Methylphosphonate mapping of phosphate contacts critical for RNA recognition by the human immunodeficiency virus tat and rev proteins

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