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

Abstract

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.

Delipidation of insect lipoprotein, lipophorin, affects its binding to the lipophorin receptor, LpR: Implications for the role of LpR-mediated endocytosis

“…In addition to the differences in density and size, the irregular, crescent-like shape of HDLp-1.17 differs from the oval shape of untreated HDLp. Such a shape was also found for HDLps of a similar density from other insect species (Ryan et al, 1990;Maya-Monteiro et al, 1997).…”

“…Using enzymatic methods, it has been shown before that the DAG and PL content of insect HDLp can be manipulated without disrupting the stability of the lipoprotein particle (Ryan et al, 1990;Kawooya et al, 1991;Rimoldi et al, 1996;Maya-Monteiro et al, 1997). Incubation of HDLp with a-cyclodextrin was shown to efficiently modulate the DAG content of HDLp from the tobacco hornworm, Manduca sexta, and the silkworm, Bombyx mori (Jouni Fig.…”

“…Importantly, however, we should not exclude the possibility that delipidated HDLp could alternatively function in delivering instead of collecting hydrophobic components other than lipids, such as hydrocarbons to the fat body at moments of food scarcity. The presence of HDLp forms of density 1.10-1.18 g/mL in the circulation has been shown to occur in M. sexta Ryan et al, 1986Ryan et al, , 1990Engler et al, 1996;Maya-Monteiro et al, 1997). In L. migratoria, by experimental alternation of food deprivation for relative short periods (12-15 h) and feeding of young adult locusts (2-4 days after imaginal ecdysis), an HDLp subspecies of density 1.13 g/mL was identified.…”

HeLp, a Heme Lipoprotein from the Hemolymph of the Cattle Tick,Boophilus microplus

“…The HeLp CD spectrum resembles those of insect lipophorins, which are characterized by a high proportion of ␤-sheet (47,48). However, despite this common feature, other aspects suggest that HeLp is a distinct particle, such as the absence of cholesterol ester in lipophorins, where diacylglycerol is the most abundant neutral lipid (49).…”

Rhodnius prolixus LIPOPHORIN: LIPID COMPOSITION AND EFFECT OF HIGH TEMPERATURE ON PHYSIOLOGICAL ROLE

“…When Manduca sexta Lp was simultaneously depleted of PLs and DAG, it precipitated (Kawooya et al, ). Dephospholipidation of R. prolixus Lp affected neither the lipoprotein structure nor its activity (Gondim et al, ; Maya‐Monteiro et al, ). On the other hand, in L. migratoria the receptor affinity for Lp depends on the lipid content (Roosendaal et al, ).…”