| Catalysis by orotidine 5'-monophosphate decarboxylase: effect of 5-fluoro and 4'-substituents on the decarboxylation of two-part substrates. | |
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MedLine Citation:
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PMID: 23276261 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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The syntheses of two novel truncated analogs of the natural substrate orotidine 5'-monophosphate (OMP) for orotidine 5'-monophosphate decarboxylase (OMPDC) with enhanced reactivity toward decarboxylation are reported: 1-(β-d-erythrofuranosyl)-5-fluoroorotic acid (FEO) and 5'-deoxy-5-fluoroorotidine (5'-dFO). A comparison of the second-order rate constants for the OMPDC-catalyzed decarboxylations of FEO (10 M⁻¹ s⁻¹) and 1-(β-d-erythrofuranosyl)orotic acid (EO, 0.026 M⁻¹ s⁻¹) shows that the vinyl carbanion-like transition state is stabilized by 3.5 kcal/mol by interactions with the 5-F substituent of FEO. The OMPDC-catalyzed decarboxylations of FEO and EO are both activated by exogenous phosphite dianion (HPO₃²⁻), but the 5-F substituent results in only a 0.8 kcal stabilization of the transition state for the phosphite-activated reaction of FEO. This provides strong evidence that the phosphite-activated OMPDC-catalyzed reaction of FEO is not limited by the chemical step of decarboxylation of the enzyme-bound substrate. Evidence is presented that there is a change in the rate-limiting step from the chemical step of decarboxylation for the phosphite-activated reaction of EO, to closure of the phosphate gripper loop and an enzyme conformational change at the ternary E•FEO•HPO₃²⁻ complex for the reaction of FEO. The 4'-CH₃ and 4'-CH₂OH groups of 5'-dFO and orotidine, respectively, result in identical destabilizations of the transition state for the unactivated decarboxylation of 2.9 kcal/mol. By contrast, the 4'-CH₃ group of 5'-dFO and the 4'-CH₂OH group of orotidine result in very different 4.7 and 8.3 kcal/mol destabilizations of the transition state for the phosphite-activated decarboxylation. Here, the destabilizing effect of the 4'-CH₃ substituent at 5'-dFO is masked by the rate-limiting conformational change that depresses the third-order rate constant for the phosphite-activated reaction of the parent substrate FEO. |
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Authors:
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Bogdana Goryanova; Krisztina Spong; Tina L Amyes; John P Richard |
Publication Detail:
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Type: Journal Article; Research Support, N.I.H., Extramural Date: 2013-01-11 |
Journal Detail:
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Title: Biochemistry Volume: 52 ISSN: 1520-4995 ISO Abbreviation: Biochemistry Publication Date: 2013 Jan |
Date Detail:
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Created Date: 2013-01-22 Completed Date: 2013-03-14 Revised Date: 2013-04-16 |
Medline Journal Info:
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Nlm Unique ID: 0370623 Medline TA: Biochemistry Country: United States |
Other Details:
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Languages: eng Pagination: 537-46 Citation Subset: IM |
Affiliation:
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Department of Chemistry, University at Buffalo, SUNY, Buffalo, NY 14260-3000, USA. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Biocatalysis Buffers Decarboxylation Enzyme Activation Enzyme Stability Kinetics Mutant Proteins / metabolism Orotic Acid / analogs & derivatives*, chemical synthesis, chemistry, metabolism Orotidine-5'-Phosphate Decarboxylase / genetics, metabolism* Osmolar Concentration Phosphites / chemistry Protein Conformation Saccharomyces cerevisiae / enzymology* Saccharomyces cerevisiae Proteins / genetics, metabolism* Substrate Specificity Uridine / analogs & derivatives, chemistry, metabolism Uridine Monophosphate / analogs & derivatives, chemistry, metabolism |
| Grant Support | |
ID/Acronym/Agency:
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GM39754/GM/NIGMS NIH HHS; R01 GM039754/GM/NIGMS NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/1-(beta-D-erythrofuranosyl)-5-fluoroorotic acid; 0/1-(beta-D-erythrofuranosyl)orotic acid; 0/5'-deoxy-5-fluoroorotidine; 0/Buffers; 0/Mutant Proteins; 0/Phosphites; 0/Saccharomyces cerevisiae Proteins; 2149-82-8/orotidylic acid; 58-96-8/Uridine; 58-97-9/Uridine Monophosphate; 65-86-1/Orotic Acid; B350MC02GZ/orotidine; EC 4.1.1.23/Orotidine-5'-Phosphate Decarboxylase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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