Effect of particle lipid content on the structure of insect lipophorins.

Ryan, Robert O.; Kay, Cyril M.; Oikawa, Kk.; Liu, H.; Bradley, R.; Scraba, Douglas G.

doi:10.1016/s0022-2275(20)41883-8

Cited by 16 publications

(5 citation statements)

References 31 publications

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“…The 45 kDa peptide was degraded after 1 h incubation with trypsin (Figure 1A). Extensive cleavage of apoLp-II was only achieved at 1 h of proteolysis, suggesting that R. prolixus apoLp-II is less exposed to the aqueous medium than apoLp-I, similarly to what has been proposed for Manduca sexta and Locusta migratoria lipophorins (Shapiro et al, 1984;Ryan et al, 1992).…”

Section: Susceptibility Of Lp To Proteolysissupporting

confidence: 70%

“…It is interesting to note that in mammalian HDL the major apoproteins are rich in surface-active amphiphilic helices (Jonas et al, 1993) which are thought to be important in stabilizing the particle in aqueous medium. For M. sexta lipophorin, it is known that the R-helix content is about 35% (Ryan et al, 1992). For R. prolixus lipophorin, analysis of the CD data (Figure 2) revealed an R-helix content of 23-27%, using the SELCON software (Sreerama & Woody, 1993).…”

Section: Discussionmentioning

confidence: 99%

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Role of Phospholipids in the Protein Stability of an Insect Lipoprotein, Lipophorin from Rhodnius prolixus

et al. 1997

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Lipophorin (Lp) is the major lipoprotein in insect hemolymph. The structural organization proposed for Lp is basically the same as that suggested for vertebrate lipoproteins, consisting of a hydrophobic core containing neutral lipids, stabilized in the aqueous environment by surrounding polar moieties of protein and phospholipids at the particle surface. After complete removal of phospholipids from Lp by phospholipase A2, the particle remains soluble [Gondim, K. C., Atella, G. C., Kawooya, J. K., & Masuda, H. (1992) Arch. Insect Biochem. Physiol. 20, 303-314]. However, studies on the roles of phospholipid on the structural stability of Lp are still lacking. In the present work, we have studied the structure and stability of dephospholipidated lipophorin (d-Lp). Trypsinolysis of d-Lp indicated no exposure of new cleavage sites on the protein when compared to Lp. However, an enhanced rate of proteolysis of the apoproteins (especially apolipophorin II) was observed in d-Lp. Circular dichroism analysis indicated that the secondary structure of Lp was not significantly affected by phospholipid removal. Furthermore, the exposure of tryptophan residues to the aqueous solvent in d-Lp was the same as in Lp, as indicated by intrinsic fluorescence emission spectra and fluorescence quenching experiments. Interestingly, d-Lp was more resistant to denaturation by guanidine hydrochloride than Lp. d-Lp was also found to be less sensitive than Lp to structural changes induced by hydrostatic pressure. Taken together, these results indicate that, although changes in its structural organization were subtle, dephospholipidated lipophorin may have additional protein-protein and/or protein-neutral lipid interactions that are responsible for the observed increase in stability. Therefore, phospholipids are not only not essential for Lp stability, but their presence in the particle seems to result in a less stable structure in the aqueous environment.

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Section: Susceptibility Of Lp To Proteolysissupporting

confidence: 70%

Section: Discussionmentioning

confidence: 99%

Role of Phospholipids in the Protein Stability of an Insect Lipoprotein, Lipophorin from Rhodnius prolixus

et al. 1997

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“…To examine the effect of apoLp-III on the PL-C dependent appearance of new DG acyl chain carbonyl resonances in HDLp, the relatively DG-deficient lipophorin subspecies, HDLp-W2, was employed in phospholipolysis experiments in the absence of exogenous apoLp-III. This subspecies, which contains ~100 DG per particle, is relatively deficient in core lipid content causing the particle to adopt an unusual, asymmetric morphology (Ryan et al" 1992). PL-C treated HDLp-W2 is relatively resistant to aggregation and the accompanying sample turbidity development, presumably because DG generated at the expense of phospholipid can be effectively stabilized by apolipophorin I and apolipophorin II (Singh et al" 1994).…”

Section: Resultsmentioning

confidence: 99%

“…On the other hand, lipophorins also have unique properties, perhaps most notably the presence of DG, not triacylglycerol and cholesteryl ester, as their major core lipid. Importantly, lipophorin particles have the capacity to accept or donate DG, forming different subspecies which have distinct compositions, structures, and morphologies (Prasad et al, 1986b;Ryan et al, 1992). During energy demanding flight, large amounts of DG are transferred to pre-existing high density lipophorin (HDLp; 300 DG molecules per particle) forming a larger, low density lipophorin (LDLp) which contains over 1000 DG molecules per particle.…”

mentioning

confidence: 99%

Identification and localization of two distinct microenvironments for the diacylglycerol component of lipophorin particles by 13C NMR

Wang¹,

Liu²,

Sykes³

et al. 1995

Biochemistry

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13C nuclear magnetic resonance spectroscopy of lipoproteins, isolated from the insect Manduca sexta, has been employed to probe the microenvironment of diacylglycerol (DG), their major neutral lipid component. Natural abundance 13C NMR spectra of high density lipophorin exhibited several well-separated resonances derived from its lipid moiety, including those for the carbonyl carbon atoms of phospholipid and DG fatty acyl chains in the region of 175-180 ppm. To verify the assignment of the DG acyl chain carbonyl carbon resonances, di[1-13C]oleoylglycerol high density lipophorin was isolated after instilling a bolus of tri[1-13C]oleoylglycerol into the midgut of larvae fed a fat-free diet. 13C NMR spectra of the isolated lipoprotein revealed a specific and dramatic enrichment of resonances at 175.5 ppm. Expansion of this region revealed two resonances separated by 0.08 ppm. These were assigned as 1,2- and 1,3- isomers of DG, the latter presumably arising from spontaneous acyl chain migration of 1,2-DG following lipoprotein isolation. On the basis of compositional and structural analysis of this lipoprotein, it is postulated that these DG species are localized predominantly in the hydrophobic core of the particle. By contrast, natural abundance 13C NMR spectra of the DG-rich, low density lipophorin (LDLp) subspecies revealed two additional resonances, separated by 0.2 ppm, that were tentatively assigned as 1,2- and 1,3-DG present at the surface of the particle. The verify this assignment, experiments employing phospholipase C, to convert lipophorin surface associated phospholipid into DG, were performed.(ABSTRACT TRUNCATED AT 250 WORDS)

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“…The same line width for phospholipid resonances within the surface monolayer of lipoproteins, however, is unknown and needs to be determined experimentally. The radius, a, of different lipophorins has been estimated from electron microscopic studies (Ryan et al, 1990;Kawooya et al, 1991;Ryan et al, 1992). LDLp and HDLp-A are spherical while…”

Section: Theorymentioning

confidence: 99%

Phosphorus-31 NMR study of the phospholipid moiety of lipophorin subspecies

et al. 1992

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31P-NMR spectra of four distinct subspecies of Manduca sexta hemolymph lipophorin revealed the presence of two resonances separated by 0.6 ppm. Phospholipid analysis of the lipoproteins showed that phosphatidylcholine (PC) and phosphatidylethanolamine (PE) were present and their mass ratio correlated well to the intensity of the two resonances in each of the different subspecies. The two resonances persisted in 31P-NMR spectra of organic solvent extracts of lipophorin. These results, together with the fact that PE, but not PC, can form an intramolecular hydrogen bond between the phosphate oxygen and the amino group of ethanolamine, resulting in deshielding of the phosphorus nucleus (and a 0.6 ppm downfield shift), strongly suggest the resonances observed represent the PC and PE components of these lipoproteins. 31P-NMR line-width data obtained as a function of temperature and solvent viscosity were used to calculate the chemical shift anisotropy (delta sigma), intrinsic viscosity (eta'), and lateral diffusion coefficients (DT) of PC and PE in different lipophorin subspecies. eta' and DT for PC and PE were similar among high-density lipophorins but differed in low-density lipophorin (LDLp). These differences may be related to the large increase in diacylglycerol content in this particle and/or the association of up to 16 molecules of apolipophorin III. On the basis of the known lipid compositional differences between LDLp and high-density lipophorin subspecies, we propose that uptake of large amounts of diacylglycerol during LDLp formation results in partitioning of this lipid to the surface monolayer where it intercalates between phospholipid molecules. Diacylglycerol intercalation creates gaps between phospholipid head groups that expose the hydrophobic surface.(ABSTRACT TRUNCATED AT 250 WORDS)

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Effect of particle lipid content on the structure of insect lipophorins.

Cited by 16 publications

References 31 publications

Role of Phospholipids in the Protein Stability of an Insect Lipoprotein, Lipophorin from Rhodnius prolixus

Role of Phospholipids in the Protein Stability of an Insect Lipoprotein, Lipophorin from Rhodnius prolixus

Identification and localization of two distinct microenvironments for the diacylglycerol component of lipophorin particles by 13C NMR

Phosphorus-31 NMR study of the phospholipid moiety of lipophorin subspecies

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