Purification and properties of a small lipid-binding protein from the hemolymph of Triatoma infestans

A small lipid-binding protein (sLBP) was purified from the hemolymph of the blood-sucking bug Triatoma infestans. Its isolation involved size exclusio n-high performance liquid chromatography (HPLC) followed by anion exchange chromatography-HPLC. The molecular weight of the protein, as determined by gel permeation chromatography, was 20 kDa. Sodium dodecyl sulphate-polyacry lamide gel electrophoresis (SDS-PAGE) resolved the protein into a single po lypeptide with M-r congruent to 16 kDa. The sLBP contains 6% lipids. Diacyl glycerols represent the major lipid class, whereas phosphatidyl-choline, ph osphatidylethanolamine, free fatty acids and hydrocarbons were found in min or amounts. The amino acid composition indicated a high content of aspartic and glutamic acids and non-polar aliphatic amino acids. The N-terminal seq uence did not resemble the sequence of any other previously reported insect hemolymph protein. Far-UV circular dichroism suggested that sLBP adopts a conformation rich in P-sheet structure. The presence of this protein in hem olymph, fat body and unfertilized eggs was explored throughout the last nym phal and adult stages of the insect by Western blot assays. These assays in dicated that sLBP is particularly abundant in hemolymph. A high concentrati on of sLBP was also detected in the fat body of the nymphs. (C) 1999 Elsevi er Science Inc. All rights reserved.