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Garcia C.F.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Cunningham M.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Soulages J.L.,Oklahoma State University | Heras H.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Garda H.A.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp
Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology | Year: 2010

Lipovitellin (LV) is essential in crustacean eggs for embryo viability and development. Two LV were isolated from eggs of Macrobrachium borellii. corresponding to early (LVe ) and late (LVl) embryo developing stages. They differ in lipid composition but not in lipid/protein ratio or apoprotein composition. Structural information was obtained by fluorescence spectroscopy, far-UV circular dichroism, partial trypsinolysis and electron microscopy applied to LVe and LVl and two partially delipidated forms of LVe generated by phospholipase A2 (LVp) or Triton X-100 (LVt) treatment. All LV forms contained two apoprotein subunits of 94 and 112 kDa, being the 112 kDa subunit more accessible to trypsinolysis in all. Only in LVp, different cleavage sites appeared. Secondary structure was similar in LVe and LVl, but LVp and LVt showed a small increase in β-sheet at expense of α-helix. Electron microscopy revealed a spheroidal morphology in all LV and a decreased size in LVp. Delipidated LVs were more resistant to denaturation with guanidinium-HCl. Acrylamide quenching of tryptophan fluorescence was more efficient in delipidated LVs, probably due to apolipoprotein rearrangement, as reinforced by fluorescence anisotropy. It is concluded that LV stability, shape, and apoprotein conformation is not largely affected by the changes in lipid composition that take place during embryogenesis.

Jaureguiberry M.S.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Jaureguiberry M.S.,National University of La Plata | Tricerri M.A.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Tricerri M.A.,National University of La Plata | And 8 more authors.
Acta Biochimica et Biophysica Sinica | Year: 2014

Experimental evidence has suggested that plasma membrane (PM)-associated signaling and hence cell metabolism and viability depend on lipid composition and organization. The aim of the present work is to develop a cell model to study the endogenous polyunsaturated fatty acids (PUFAs) effect on PM properties and analyze its influence on cholesterol (Chol) homeostasis. We have previously shown that by using a cell line over-expressing stearoyl-CoA-desaturase, membrane composition and organization coordinate cellular pathways involved in Chol efflux and cell viability by different mechanisms. Now, we expanded our studies to a cell model over-expressing both Δ5 and Δ6 desaturases, which resulted in a permanently higher PUFA content in PM. Furthermore, this cell line showed increased PM fluidity, Chol storage, and mitochondrial activity. In addition, human apolipoprotein A-I-mediated Chol removal was less efficient in these cells than in the corresponding control. Taken together, our results suggested that the cell functionality is preserved by regulating PM organization and Chol exportation and homeostasis. © 2014 © The Author 2014. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

Cuellar L.A.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Prieto E.D.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Cabaleiro L.V.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp | Garda H.A.,Institute Investigaciones Bioquimicas Of La Plata Inibiolp
Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids | Year: 2014

Discoidal high-density lipoproteins (D-HDL) are critical intermediates in reverse cholesterol transport. Most of the present knowledge of D-HDL is based on studies with reconstituted lipoprotein complexes of apolipoprotein A-I (apoA-I) obtained by cholate dialysis (CD). D-HDL can also be generated by the direct microsolubilization (DM) of phospholipid vesicles at the gel/fluid phase transition temperature, a process mechanistically similar to the "in vivo" apoAI lipidation via ABCA1. We compared the apoA-I configuration in D-HDL reconstituted with dimyristoylphosphatidylcholine by both procedures using fluorescence resonance energy transfer measurements with apoA-I tryptophan mutants and fluorescently labeled cysteine mutants. Results indicate that apoA-I configuration in D-HDL depends on the reconstitution process and are consistent with a "double belt" molecular arrangement with different helix registry. As reported by others, a configuration with juxtaposition of helices 5 of each apoAI monomer (5/5 registry) predominates in D-HDL obtained by CD. However, a configuration with helix 5 of one monomer juxtaposed with helix 2 of the other (5/2 registry) would predominate in D-HDL generated by DM. Moreover, we also show that the kinetics of cholesterol efflux from macrophage cultures depends on the reconstitution process, suggesting that apoAI configuration is important for this HDL function. © 2013 Elsevier B.V.

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