Document Detail


Measurement of microsecond dynamic motion in the intestinal fatty acid binding protein by using fluorescence correlation spectroscopy.
MedLine Citation:
PMID:  12381795     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Fluorescence correlation spectroscopy (FCS) measurements have been carried out on the intestinal fatty acid binding protein (IFABP) to study microsecond dynamics of the protein in its native state as well as in pH-induced intermediates. IFABP is a small (15 kDa) protein that consists mostly of antiparallel beta-strands enclosing a large central cavity into which the ligand binds. Because this protein does not contain cysteine, two cysteine mutants (Val60Cys and Phe62Cys) have been prepared and covalently modified with fluorescein. Based on fluorescence measurements, one of the mutants (Val60Flu) has the fluorescein moiety inside the cavity of the protein, whereas the fluorescein is exposed to solvent in the other (Phe62Flu). The protein modified at position 60 demonstrates the presence of a conformational event on the order of 35 microsec, which is not seen in the other mutant (Phe62Flu). The amplitude of this fast conformational event decreases sharply at low pH as the protein unfolds. Experiments measuring the diffusion as a function of pH indicate the formation of a compact state distinct from the native state at about pH 3.5. Steady state fluorescence and far-UV CD indicates that unfolding occurs at pH values below pH 3.
Authors:
Krishnananda Chattopadhyay; Saveez Saffarian; Elliot L Elson; Carl Frieden
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.     Date:  2002-10-15
Journal Detail:
Title:  Proceedings of the National Academy of Sciences of the United States of America     Volume:  99     ISSN:  0027-8424     ISO Abbreviation:  Proc. Natl. Acad. Sci. U.S.A.     Publication Date:  2002 Oct 
Date Detail:
Created Date:  2002-10-30     Completed Date:  2002-12-09     Revised Date:  2009-11-18    
Medline Journal Info:
Nlm Unique ID:  7505876     Medline TA:  Proc Natl Acad Sci U S A     Country:  United States    
Other Details:
Languages:  eng     Pagination:  14171-6     Citation Subset:  IM    
Affiliation:
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO 63110, USA.
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MeSH Terms
Descriptor/Qualifier:
Carrier Proteins / analysis*,  chemistry,  genetics
Diffusion
Fatty Acid-Binding Proteins
Fluorescence
Hydrogen-Ion Concentration
Isomerism
Neoplasm Proteins*
Protein Folding*
Spectrometry, Fluorescence / methods
Grant Support
ID/Acronym/Agency:
DK13332/DK/NIDDK NIH HHS; GM38838/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Carrier Proteins; 0/Fatty Acid-Binding Proteins; 0/Neoplasm Proteins
Comments/Corrections

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