Deletion of the propeptide of apolipoprotein A-I reduces protein expression but stimulates effective conversion of preβ-high density lipoprotein to α-high density lipoprotein

Sviridov, Dmitri; Pyle, Louise E.; Jauhiainen, Matti; Ehnholm, Christian; Fidge, Noel

doi:10.1016/s0022-2275(20)31981-7

Cited by 17 publications

(3 citation statements)

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“…We (4,5) and others (6) have demonstrated that expression of recombinant apoA-I with the pro-peptide deleted impairs secretion and decreases the overall efficiency of the expression of apoA-I, suggesting that apoA-I is synthesized as a pro-protein to aid in intracellular transport of the protein through the cellular apparatus. We have also demonstrated that the presence of pro-peptide impairs interconversion of preβ-HDL into R-HDL (5), suggesting a requirement for its cleavage. An apoA-I(P3R) substitution has been recorded, in which the individual was found to have a proapoA-I concentration of the variant protein approximately 3 times the level of the normal protein; however, no detectable disease state was recognizable with this mutation (7).…”

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confidence: 92%

“…The functional role of the pro-peptide, the requirement for its subsequent cleavage, and the structural features essential for this cleavage are unclear. We ( , ) and others () have demonstrated that expression of recombinant apoA-I with the pro-peptide deleted impairs secretion and decreases the overall efficiency of the expression of apoA-I, suggesting that apoA-I is synthesized as a pro-protein to aid in intracellular transport of the protein through the cellular apparatus. We have also demonstrated that the presence of pro-peptide impairs interconversion of preβ-HDL into α-HDL (), suggesting a requirement for its cleavage.…”

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confidence: 92%

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Characterization of the Maturation of Human Pro-apolipoprotein A-I in an in Vitro Model

2001

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The reaction conditions and the protein structural features involved in the maturation of pro-apolipoprotein A-I (cleavage of pro-peptide) were investigated in an in vitro model. ProapoA-I, mutants and wild type, were expressed in the PGEX/E. coli expression system as fusion proteins with glutathione S-transferase (GST). Use of GST-proapoA-I and truncated forms of proapoA-I enabled quantitation of the amount of GST and apoA-I formed as a result of cleavage following incubation with human serum. Deletion of the pro-peptide (GST-apoA-I) resulted in complete inhibition of the reaction. Truncation of proapoA-I to residues 222, 150, 135, and 25 as well as substitution of residues -6, -5, and -4 with alanine did not affect the reaction. Substitution of residues -1, -2, 1, 3, and 4 with alanine either completely blocked or substantially inhibited cleavage of the pro-peptide. The reaction was inhibited by addition of EDTA, o-phenanthroline, dithiothreitol, and beta-mercaptoethanol and to a lesser extent by p-chloromercuriphenylsulfonic acid, but not by leupeptin, N-ethylmaleimide, PMSF, pepstatin A, or trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane. Calcium was essential for the activation of the cleavage enzyme, but it had a biphasic effect on the cleavage, activating it at concentrations below 1.5 mM and inhibiting at concentrations above 1.75 mM. Manganese alone was not essential for activation of the enzyme nor did it modify the effect of low concentration of calcium. However, a high concentration of manganese partially reverted the inhibitory effect of a high calcium concentration. Thus, residues within -2 to +4 are involved in forming the cleavage site for the maturation enzyme. The reaction of maturation is inhibited by metalloprotease inhibitors and is dependent upon calcium.

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confidence: 92%

Characterization of the Maturation of Human Pro-apolipoprotein A-I in an in Vitro Model

2001

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“…LDL was purified from human plasma by sequential centrifugation and acetylated as described by Basu et al [53]. Cholesterol and phospholipid efflux were assessed as described previously [54]. Briefly, fibroblasts were incubated in labeling medium containing [ 3 H]cholesterol (75 kBq/ml) or [methyl- 14 C] choline (0.2 MBq/ml) for 48 hours.…”

Section: Lipid Efflux Assaysmentioning

confidence: 99%

Correction: A Mouse Model of Harlequin Ichthyosis Delineates a Key Role for Abca12 in Lipid Homeostasis

Smyth¹,

Hacking²,

Hilton³

et al. 2008

PLoS Genet

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Harlequin Ichthyosis (HI) is a severe and often lethal hyperkeratotic skin disease caused by mutations in the ABCA12 transport protein. In keratinocytes, ABCA12 is thought to regulate the transfer of lipids into small intracellular trafficking vesicles known as lamellar bodies. However, the nature and scope of this regulation remains unclear. As part of an original recessive mouse ENU mutagenesis screen, we have identified and characterised an animal model of HI and showed that it displays many of the hallmarks of the disease including hyperkeratosis, loss of barrier function, and defects in lipid homeostasis. We have used this model to follow disease progression in utero and present evidence that loss of Abca12 function leads to premature differentiation of basal keratinocytes. A comprehensive analysis of lipid levels in mutant epidermis demonstrated profound defects in lipid homeostasis, illustrating for the first time the extent to which Abca12 plays a pivotal role in maintaining lipid balance in the skin. To further investigate the scope of Abca12's activity, we have utilised cells from the mutant mouse to ascribe direct transport functions to the protein and, in doing so, we demonstrate activities independent of its role in lamellar body function. These cells have severely impaired lipid efflux leading to intracellular accumulation of neutral lipids. Furthermore, we identify Abca12 as a mediator of Abca1-regulated cellular cholesterol efflux, a finding that may have significant implications for other diseases of lipid metabolism and homeostasis, including atherosclerosis.

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Concentration and pattern changes of porcine serum apolipoprotein A‐I in four different infectious diseases

et al. 2015

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Apolipoprotein A-I (Apo A-I) is a major protein in lipid/lipoprotein metabolism and decreased serum levels have been observed in many species in response to inflammatory and infectious challenges. Little is known about the porcine homologue, therefore in this work we have characterized it through biochemical and proteomic techniques. In 2DE, porcine serum Apo A-I is found as three spots, the two more acidic ones corresponding to the mature protein, the more basic spot to the protein precursor. Despite high sequence coverage in LC-MS/MS, we did not find a sequence or PTM difference between the two mature protein species. Besides this biochemical characterization, we measured overall levels and relative species abundance of serum Apo A-I in four different viral and bacterial porcine infectious diseases. Lower overall amounts of Apo A-I were observed in Salmonella typhimurium and Escherichia coli infections. In the 2DE protein pattern, an increase of the protein precursor together with a lower level of mature protein species were detected in the porcine circovirus type 2-systemic disease and S. typhimurium infection. These results reveal that both the porcine serum Apo A-I concentration and the species pattern are influenced by the nature of the infectious disease.

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Deletion of the propeptide of apolipoprotein A-I reduces protein expression but stimulates effective conversion of preβ-high density lipoprotein to α-high density lipoprotein

Cited by 17 publications

References 56 publications

Characterization of the Maturation of Human Pro-apolipoprotein A-I in an in Vitro Model

Characterization of the Maturation of Human Pro-apolipoprotein A-I in an in Vitro Model

Correction: A Mouse Model of Harlequin Ichthyosis Delineates a Key Role for Abca12 in Lipid Homeostasis

Concentration and pattern changes of porcine serum apolipoprotein A‐I in four different infectious diseases

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