Functional and Structural Characterization of a Synthetic Peptide Representing the N-Terminal Domain of Prokaryotic Pyruvate Dehydrogenase

A synthetic peptide (Nterm-E1p) is used to characterize the structure and function of the N-terminal region (amino acid residues 4−45) of the pyruvate dehydrogenase component (E1p) from the pyruvate dehydrogenase multienzyme complex (PDHC) from Azotobacter vinelandii. Activity and binding studies es...

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Bibliographic Details
Published in:Biochemistry (Easton) 2002-06, Vol.41 (23), p.7490-7500
Main Authors: Hengeveld, Annechien F, van Mierlo, Carlo P. M, van den Hooven, Henno W, Visser, Antonie J. W. G, de Kok, Aart
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Azotobacter vinelandii - enzymology
Biochemie
Biochemistry
Dimerization
Guanidine - chemistry
Hydrogen-Ion Concentration
Models, Molecular
Molecular Sequence Data
Molecular Weight
Nuclear Magnetic Resonance, Biomolecular
Peptide Fragments - chemical synthesis
Peptide Fragments - chemistry
Peptide Fragments - physiology
Protein Conformation
Protein Denaturation
Protein Folding
Protein Structure, Secondary
Protein Structure, Tertiary
Pyruvate Dehydrogenase Complex - chemical synthesis
Pyruvate Dehydrogenase Complex - chemistry
Pyruvate Dehydrogenase Complex - physiology
Spectrometry, Fluorescence
Structure-Activity Relationship
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Summary:A synthetic peptide (Nterm-E1p) is used to characterize the structure and function of the N-terminal region (amino acid residues 4−45) of the pyruvate dehydrogenase component (E1p) from the pyruvate dehydrogenase multienzyme complex (PDHC) from Azotobacter vinelandii. Activity and binding studies established that Nterm-E1p specifically competes with E1p for binding to the dihydrolipoyl transacetylase component (E2p) of PDHC. Moreover, the experiments show that the N-terminal region of E1p forms an independent folding domain that functions as a binding domain. CD measurements, two-dimensional (2D) 1H NMR analysis, and secondary structure prediction all indicate that Nterm-E1p has a high α-helical content. Here a structural model of the N-terminal domain is proposed. The peptide is present in two conformations, the population of which depends on the sample conditions. The conformations are designated “unfolded” at pH ≥6 and “folded” at pH
ISSN:0006-2960
1520-4995
DOI:10.1021/bi012172u