Secretion of a novel, developmentally regulated fatty acid-binding protein into the perivitelline fluid of the parasitic nematode, Ascaris suum.
Early development of the parasitic nematode, Ascaris suum, occurs inside a highly resistant eggshell, and the developing larva is bathed in perivitelline fluid. Two-dimensional gel analysis of perivitelline fluid from infective larvae reveals seven major proteins; a cDNA encoding one of these, As-p18, has been cloned, sequenced, and protein expressed in Escherichia coli. The predicted amino acid sequence of As-p18 exhibits similarities to the intracellular lipid-binding protein (iLBP) family including retinoid- and fatty acid-binding proteins (FABP). As-p18 is unusual in that it possesses a hydrophobic leader that is not present in the mature protein, the developmental regulation of its expression, and in terms of its predicted structure. Recombinant As-p18 is a functional FABP with a high affinity for both a fluorescent fatty acid analog (11(((5-(dimethylamino)-1-naphthalenyl)sulfonyl)amino) undecanoic acid) and oleic acid, but not retinol. Circular dichroism of rAs-p18 reveals a high beta-sheet content (62%), which is consistent with secondary structure for the protein predicted from sequence algorithms, and the structure of iLBPs. Unusual features are apparent in a structural model of As-p18 generated from existing crystal structures of iLBPs. As-p18 is not found in unembryonated eggs, begins to be synthesized at about day 3 of development, reaches a maximal concentration with the formation of the first-stage larva and remains abundant in the perivitelline fluid of the second-stage larva. Since As-p18 is not present in the post-infective third-stage larva or adult worm tissues, it appears to be exclusive to the egg. Surprisingly, however, Northern blot analysis yields mRNA for As-p18 not only in the early larval stages, but also the unembryonated egg, third-stage larvae, and ovaries of adult worms, even though the protein is not detectable from any of those sources. As-p18 may play a role in sequestering potentially toxic fatty acids and their peroxidation products, or it may be involved in the maintenance of the impermeable lipid layer of the eggshell.
Mei
B
B
Department of Biology, The University of Toledo, Toledo, Ohio 43606-3390, USA.
Kennedy
M W
MW
Beauchamp
J
J
Komuniecki
P R
PR
Komuniecki
R
R
D000595:Amino Acid Sequence; D000818:Animals; D017165:Ascaris suum; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002942:Circular Dichroism; D015180:Electrophoresis, Gel, Two-Dimensional; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids / Q000378:metabolism*; D015801:Helminth Proteins; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018996:Myelin P2 Protein / Q000737:chemistry / Q000378:metabolism*; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D011487:Protein Conformation; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014817:Vitelline Membrane / Q000737:chemistry*
C105516:As-p18 protein, Ascaris suum; D002352:Carrier Proteins; C082667:Fabp5 protein, mouse; C086936:Fabp7 protein, mouse; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D015801:Helminth Proteins; D018996:Myelin P2 Protein; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins