The DvA-1 polyprotein of the parasitic nematode Dictyocaulus viviparus. A small helix-rich lipid-binding protein.
DNA encoding a single unit of the DvA-1 polyprotein of the parasitic nematode Dictyocaulus viviparus was isolated and the polypeptide ("rDvA-1L") expressed in Escherichia coli, to give a protein showing high binding affinity for fatty acids and retinoids. Fluorescent fatty acid probes show substantial changes in emission spectrum in the presence of rDvA-1L, which can be reversed by fatty acids (oleic, palmitic, stearic, arachidonic) and retinoids, but not by tryptophan, squalene, or cholesterol. Moreover, changes in intrinsic fluorescence of retinol or retinoic acid confirm a retinoid binding activity. Fluorescence titration experiments indicate stoichiometric binding to a single protein site per monomer unit with affinities (Kd) in the range 3 x 10(-8) M for 11-((5-dimethylaminonaphthalene-1-sulfonyl)amino)undecanoic acid, and by competition, 5 x 10(-8) M for oleic acid. The extreme blue shift of bound fluorescent fatty acid suggests an unusually low polarity for the protein binding site. The emission spectrum of the single tryptophan of rDvA-1L indicates that it is deeply buried in a nonpolar environment, and its spectrum is unaffected by ligand binding. Far UV circular dichroism of rDvA-1L reveals a high alpha-helix content (53%). Differential scanning calorimetry studies indicate that rDvA-1L is highly stable (T(m) approximately 98 degrees C), refolding efficiently following thermal denaturation. DvA-1 therefore represents an example of a new class of lipid binding protein, and is the first product of a polyprotein with this activity to be described.
Kennedy
M W
MW
Wellcome Laboratories for Experimental Parasitology, Glasgow, Scotland, United Kingdom.
Britton
C
C
Price
N C
NC
Kelly
S M
SM
Cooper
A
A
D000818:Animals; D001483:Base Sequence; D002152:Calorimetry, Differential Scanning; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D002942:Circular Dichroism; D017931:DNA Primers; D004021:Dictyocaulus / Q000737:chemistry*; D050556:Fatty Acid-Binding Proteins; D015801:Helminth Proteins / Q000737:chemistry* / Q000378:metabolism; D050356:Lipid Metabolism; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
D002352:Carrier Proteins; D017931:DNA Primers; D050556:Fatty Acid-Binding Proteins; D015801:Helminth Proteins; D009363:Neoplasm Proteins