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| | {{STRUCTURE_1jn4| PDB=1jn4 | SCENE= }} | | {{STRUCTURE_1jn4| PDB=1jn4 | SCENE= }} |
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| - | '''The Crystal Structure of Ribonuclease A in complex with 2'-deoxyuridine 3'-pyrophosphate (P'-5') adenosine'''
| + | ===The Crystal Structure of Ribonuclease A in complex with 2'-deoxyuridine 3'-pyrophosphate (P'-5') adenosine=== |
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| - | ==Overview==
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| - | Recently, 3',5'-pyrophosphate-linked 2'-deoxyribodinucleotides were shown to be >100-fold more effective inhibitors of RNase A superfamily enzymes than were the corresponding monophosphate-linked (i.e., standard) dinucleotides. Here, we have investigated two ribo analogues of these compounds, cytidine 3'-pyrophosphate (P'-->5') adenosine (CppA) and uridine 3'-pyrophosphate (P'-->5') adenosine (UppA), as potential substrates for RNase A and angiogenin. CppA and UppA are cleaved efficiently by RNase A, yielding as products 5'-AMP and cytidine or uridine cyclic 2',3'-phosphate. The k(cat)/K(m) values are only 4-fold smaller than for the standard dinucleotides CpA and UpA, and the K(m) values (10-16 microM) are lower than those reported for any earlier small substrates (e.g., 500-700 microM for CpA and UpA). The k(cat)/K(m) value for CppA with angiogenin is also only severalfold smaller than for CpA, but the effect of lengthening the internucleotide linkage on K(m) is more modest. Ribonucleotide 3',5'-pyrophosphate linkages were proposed previously to exist in nature as chemically labile intermediates in the pathway for the generation of cyclic 2',3'-phosphate termini in various RNAs. We demonstrate that in fact they are relatively stable (t(1/2) > 15 days for uncatalyzed degradation of UppA at pH 6 and 25 degrees C) and that cleavage in vivo is most likely enzymatic. Replacements of the RNase A catalytic residues His12 and His119 by alanine reduce activity toward UppA by approximately 10(5)-and 10(3.3)-fold, respectively. Thus, both residues play important roles. His12 probably acts as a base catalyst in cleavage of UppA (as with RNA). However, the major function of His119 in RNA cleavage, protonation of the 5'-O leaving group, is not required for UppA cleavage because the pK(a) of the leaving group is much lower than that for RNA substrates. A crystal structure of the complex of RNase A with 2'-deoxyuridine 3'-pyrophosphate (P'-->5') adenosine (dUppA), determined at 1.7 A resolution, together with models of the UppA complex based on this structure suggest that His119 contributes to UppA cleavage through a hydrogen bond with a nonbridging oxygen atom in the pyrophosphate and through pi-pi stacking with the six-membered ring of adenine.
| + | The line below this paragraph, {{ABSTRACT_PUBMED_11513604}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 11513604 is the PubMed ID number. |
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| | + | {{ABSTRACT_PUBMED_11513604}} |
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| | ==About this Structure== | | ==About this Structure== |
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| | [[Category: Nucleotide inhibitor]] | | [[Category: Nucleotide inhibitor]] |
| | [[Category: Ribonuclease]] | | [[Category: Ribonuclease]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 21:26:35 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Tue Jul 1 20:29:50 2008'' |
Revision as of 17:29, 1 July 2008
Template:STRUCTURE 1jn4
The Crystal Structure of Ribonuclease A in complex with 2'-deoxyuridine 3'-pyrophosphate (P'-5') adenosine
Template:ABSTRACT PUBMED 11513604
About this Structure
1JN4 is a Single protein structure of sequence from Bos taurus. Full crystallographic information is available from OCA.
Reference
Cleavage of 3',5'-pyrophosphate-linked dinucleotides by ribonuclease A and angiogenin., Jardine AM, Leonidas DD, Jenkins JL, Park C, Raines RT, Acharya KR, Shapiro R, Biochemistry. 2001 Aug 28;40(34):10262-72. PMID:11513604
Page seeded by OCA on Tue Jul 1 20:29:50 2008
