دورية أكاديمية
Hemiacetal stabilization in a chymotrypsin inhibitor complex and the reactivity of the hydroxyl group of the catalytic serine residue of chymotrypsin.
العنوان: | Hemiacetal stabilization in a chymotrypsin inhibitor complex and the reactivity of the hydroxyl group of the catalytic serine residue of chymotrypsin. |
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المؤلفون: | Cleary JA; School of Biomolecular and Biomedical Science, Centre for Synthesis and Chemical Biology, Conway Institute, University College Dublin, Dublin 4, Ireland., Doherty W; School of Chemistry and Chemical Biology, Centre for Synthesis and Chemical Biology, Conway Institute, University College Dublin, Dublin 4, Ireland., Evans P; School of Chemistry and Chemical Biology, Centre for Synthesis and Chemical Biology, Conway Institute, University College Dublin, Dublin 4, Ireland., Malthouse JP; School of Biomolecular and Biomedical Science, Centre for Synthesis and Chemical Biology, Conway Institute, University College Dublin, Dublin 4, Ireland. Electronic address: J.Paul.G.Malthouse@ucd.ie. |
المصدر: | Biochimica et biophysica acta [Biochim Biophys Acta] 2014 Jun; Vol. 1844 (6), pp. 1119-27. Date of Electronic Publication: 2014 Mar 21. |
نوع المنشور: | Journal Article; Research Support, Non-U.S. Gov't |
اللغة: | English |
بيانات الدورية: | Publisher: Elsevier Pub. Co Country of Publication: Netherlands NLM ID: 0217513 Publication Model: Print-Electronic Cited Medium: Print ISSN: 0006-3002 (Print) Linking ISSN: 00063002 NLM ISO Abbreviation: Biochim Biophys Acta Subsets: MEDLINE |
أسماء مطبوعة: | Original Publication: Amsterdam : Elsevier Pub. Co. |
مواضيع طبية MeSH: | Chymotrypsin/*chemistry , Histidine/*chemistry , Oligopeptides/*chemistry , Protease Inhibitors/*chemistry , Serine/*chemistry, Animals ; Biocatalysis ; Catalytic Domain ; Cattle ; Glyoxal/chemistry ; Hydrogen Bonding ; Hydrogen-Ion Concentration ; Kinetics ; Nuclear Magnetic Resonance, Biomolecular ; Oligopeptides/chemical synthesis ; Protease Inhibitors/chemical synthesis ; Solutions ; Stereoisomerism ; Thermodynamics |
مستخلص: | The aldehyde inhibitor Z-Ala-Ala-Phe-CHO has been synthesized and shown by (13)C-NMR to react with the active site serine hydroxyl group of alpha-chymotrypsin to form two diastereomeric hemiacetals. For both hemiacetals oxyanion formation occurs with a pKa value of ~7 showing that chymotrypsin reduces the oxyanion pKa values by ~5.6 pKa units and stabilizes the oxyanions of both diastereoisomers by ~32kJmol(-1). As pH has only a small effect on binding we conclude that oxyanion formation does not have a significant effect on binding the aldehyde inhibitor. By comparing the binding of Z-Ala-Ala-Phe-CHO with that of Z-Ala-Ala-Phe-H we estimate that the aldehyde group increases binding ~100 fold. At pH7.2 the effective molarity of the active site serine hydroxy group is ~6000 which is ~7× less effective than with the corresponding glyoxal inhibitor. Using (1)H-NMR we have shown that at both 4 and 25°C the histidine pKa is ~7.3 in free chymotrypsin and it is raised to ~8 when Z-Ala-Ala-Phe-CHO is bound. We conclude that oxyanion formation only has a minor role in raising the histidine pKa and that the aldehyde hydrogen must be replaced by a larger group to raise the histidine pKa>10 and give stereospecific formation of tetrahedral intermediates. The results show that a large increase in the pKa of the active site histidine is not needed for the active site serine hydroxyl group to have an effective molarity of 6000. (Copyright © 2014 Elsevier B.V. All rights reserved.) |
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معلومات مُعتمدة: | United Kingdom Wellcome Trust; 055637/Z/98 United Kingdom WT_ Wellcome Trust |
فهرسة مساهمة: | Keywords: Aldehyde inhibitor; Chymotrypsin; Effective molarity; Hemiacetal; Oxyanion |
المشرفين على المادة: | 0 (Oligopeptides) 0 (Protease Inhibitors) 0 (Solutions) 452VLY9402 (Serine) 4QD397987E (Histidine) 50NP6JJ975 (Glyoxal) EC 3.4.21.1 (Chymotrypsin) EC 3.4.21.1 (alpha-chymotrypsin) |
تواريخ الأحداث: | Date Created: 20140325 Date Completed: 20140626 Latest Revision: 20220216 |
رمز التحديث: | 20221213 |
مُعرف محوري في PubMed: | PMC7185751 |
DOI: | 10.1016/j.bbapap.2014.03.008 |
PMID: | 24657307 |
قاعدة البيانات: | MEDLINE |
تدمد: | 0006-3002 |
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DOI: | 10.1016/j.bbapap.2014.03.008 |