Document Detail


Potentiometric determination of ionizations at the active site of papain.
MedLine Citation:
PMID:  10964     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The ionization behavior of groups at the active site of papain was determined from the pH dependence of the difference of proton content of papain and the methylthio derivative of the thiol group at the active site of papain (papain-S-SCH3). This difference in proton content was determined directly by two independent methods. One method involved potentiometric measurements of the protons released and demethylthiolation of papain-S-SCH3 with dithiothreitol, as a function of pH. The other method involved analogous measurements of the protons released on methylthiolation of papain with methyl methanethiosulfonate. The methylthio pH-difference titrations generated by these measurements indicate that ionization of the thiol group at the active site of papain is linked to the ionization of His-159. The pK of the thiol group changes from 3.3 to 7.6 on deprotonation of His-159 at 29 degrees C/20.05. Similarly, the pK of His-159 shifts from 4.3 to 8.5 when the active site thiol group is deprotonated. The microscopic ionization constants determined in this work for Cys-25 and His-159 indicate that equilibrium constant for transfer of the proton from Cys-25 to His-159 is 8--12, and that in the physiological pH range the active site thiol group exists mainly as a thiol anion.
Authors:
S D Lewis; F A Johnson; J A Shafer
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biochemistry     Volume:  15     ISSN:  0006-2960     ISO Abbreviation:  Biochemistry     Publication Date:  1976 Nov 
Date Detail:
Created Date:  1977-01-29     Completed Date:  1977-01-29     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  5009-17     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Binding Sites
Dithiothreitol / pharmacology
Hydrogen-Ion Concentration
Mathematics
Papain* / metabolism
Potentiometry
Protein Binding
Spectrometry, Fluorescence
Thermodynamics
Chemical
Reg. No./Substance:
3483-12-3/Dithiothreitol; EC 3.4.22.2/Papain

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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