کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10536917 | 962635 | 2014 | 8 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Energy cost for the proper ionization of active site residues in 6-phosphogluconate dehydrogenase from T. brucei
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کلمات کلیدی
ITCDTNBTrypanosoma brucei6PGDHIonization enthalpyRU5P6PG6-phosphogluconate dehydrogenase - 6-فسفوگلاوکونات دهیدروژناز6-phosphogluconate - 6-فسفوگلوکوناتtriethanolamine hydrochloride - تری اتانولامین هیدروکلرایدribulose-5-phosphate - ریوبولوز 5-فسفاتwild type - نوع وحشیProton linkage - پیوند پروتونTEA - چایIsothermal titration calorimetry - کالری سنجی تیتاسیون ایزوترمال
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
The catalytic mechanism of 6-phosphogluconate dehydrogenase requires the inversion of a Lys/Glu couple from its natural ionization state. The pKa of these residues in free and substrate bound enzymes has been determined measuring by ITC the proton release/uptake induced by substrate binding at different pH values. Wt 6-phosphogluconate dehydrogenase from Trypanosoma brucei and two active site enzyme mutants, K185H and E192Q were investigated. Substrate binding was accompanied by proton release and was dependent on the ionization of a group with pKa 7.07 which was absent in the E192Q mutant. Kinetic data highlighted two pKa, 7.17 and 9.64, in the enzyme-substrate complex, the latter being absent in the E192Q mutant, suggesting that the substrate binding shifts Glu192 pKa from 7.07 to 9.64. A comparison of wt and E192Q mutant appears to show that the substrate binding shifts Lys185 pKa from 9.9 to 7.17. By comparing differences in proton release and the binding enthalpy of wt and mutant enzymes, the enthalpic cost of the change in the protonation state of Lys185 and Glu192 was estimated at â 6.1 kcal/mol. The change in protonation state of Lys185 and Glu192 has little effect on Gibbs free energy, 240-325 cal/mol. However proton balance evidences the dissociation of other group(s) that can be collectively described by a single pKa shift from 9.1 to 7.54. This further change in ionization state of the enzyme causes an increase of free energy with a total cost of 1.2-2.3 kcal/mol to set the enzyme into a catalytically competent form.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics - Volume 1844, Issue 4, April 2014, Pages 785-792
Journal: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics - Volume 1844, Issue 4, April 2014, Pages 785-792
نویسندگان
S. Hanau, L. Proietti d'Empaire, K. Montin, C. Cervellati, I. Capone, F. Dallocchio,