[31] Immunochemical methods for studying lipoprotein structure

Krul, Elaine S.; Schonfeld, Gustav

doi:10.1016/0076-6879(86)28091-x

Cited by 9 publications

(4 citation statements)

References 78 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although a more conclusive proof would require studies of site-directed mutagenesis, the location of the thyroid hormone domain is consistent with a number of independent data (2)(3)(4)(5)(6). For instance, in apo A-I the position of the epitope (aa 1-16) of the MAb that inhibited T4 binding with the greatest potency overlapped the position of the region (aa [8][9][10][11][12][13][14][15][16][17][18][19][20][21][22] where the homology with the other exon 3-coded HDL apolipoproteins, as well as with TBG, TTR, and HSA, is maximal (4) and also overlapped the position (aa 9-23) of the computer-predicted highest beta sheet potential (2). The T4 binding sites of TBG and TTR are in regions having beta sheet structure (2,3).…”

supporting

confidence: 60%

See 1 more Smart Citation

A Thyroid Hormone Binding Motif Is Evolutionarily Conserved in Apolipoproteins

Benvenga

1997

Thyroid

View full text Add to dashboard Cite

High-density lipoproteins (HDL) are the major lipoprotein (Lp) plasma carriers of thyroid hormones, binding mediated by a specific interaction with their apolipoproteins (A-I, A-II, A-IV, C-I, C-II, C-III and E). The single binding site of these apolipoproteins is encoded by exon 3 (exon 2 for apoA-IV) of the respective gene and has amino acid sequence homology with regions of the three major thyroid hormone plasma transport proteins (TBG, TTR, and albumin) known to contain the corresponding hormone binding site(s). Within the hormone domain, we identified a 5-residue hydrophobic motif "Y, L/I/M, X, X, V/L/I" that is extremely well conserved in apolipoproteins. The exon-3 coded region of human apo E contains a hydrophobic pocket which is formed by Trp 34 and a number of neighboring leucines (residues no. 28, 30, 37 or 43). The location of this pocket overlaps strikingly that of the region (aa 26-40) where the said homology is maximal and where the motif YLRVW, (aa 36-40) lies. This hydrophobic pocket should represent the thyroid hormone site of apo E and, because of the said homology, should exist in the other HDL apolipoproteins. Because TBG and/or TTR are not present in all animal species, but Lp are, and because fish HDL bind thyroid hormones, I postulated that thyroid hormone binding to HDL apolipoproteins is conserved through the phylum. To this end, I evaluated the conservation of the 5-residue motif in all the apolipoprotein sequences known (PIR data bank no. 42) and tested the thyroid hormone binding properties of two animal apolipoproteins that were available (bovine apo A-I and rabbit apo E). I found that the conservation does exist and that the binding properties of the two animal apolipoproteins match those of the respective human counterpart. In addition, I found that the 5-residue motif is spared by naturally occurring mutations, which is not the case for other domains. I therefore conclude that the interaction of thyroid hormones with Lp represents the first plasma transport for these hormones that appeared in the animal world and that preservation of the structure of the hormone domain appears to be more important than preservation of other domains.

show abstract

supporting

confidence: 60%

“…Indeed, there is an increasing use of MAb as invaluable tools to probe the physical and functional properties of proteins, including apolipoproteins (8). Although a more conclusive proof would require studies of site-directed mutagenesis, the location of the thyroid hormone domain is consistent with a number of independent data (2)(3)(4)(5)(6).…”

mentioning

confidence: 72%

A Thyroid Hormone Binding Motif Is Evolutionarily Conserved in Apolipoproteins

Benvenga

1997

Thyroid

View full text Add to dashboard Cite

show abstract

“…Although speculative, during HDL particle assembly PON may associate with the forming HDL in a series of interactions. It is known that, upon lipidation, apoA-I undergoes conformational changes detectable with secondary structure-specific monoclonal antibodies ( − ). Because the structure of lipid-free apoA-I is significantly different than that of lipidated apoA-I, it is reasonable to speculate that, during lipidation, regions of apoA-I become transiently available for PON interaction.…”

Section: Discussionmentioning

confidence: 99%

Cysteine Substitutions in Apolipoprotein A-I Primary Structure Modulate Paraoxonase Activity

et al. 2001

View full text Add to dashboard Cite

Paraoxonase (PON) is transported primarily on apolipoprotein A-I (apoA-I) -containing highdensity lipoprotein (HDL) and is thought to protect against early atherogenic events including low-density lipoprotein (LDL) oxidation and monocyte migration. It has been proposed that apoA-I may be necessary for PON's association with plasma HDL. On the basis of this, we examined the effect of apoA-I on PON's enzymatic activity and its ability to associate with HDL. Additionally, we examined whether changes in apoA-I primary structure (cysteine substitution mutations) could modulate these effects. Chinese hamster ovary cells stably transfected with human PON1A cDNA were incubated in the presence and absence of recombinant wild-type apoA-I (apoA-I WT ) and specific Cys substitution mutations. Extracellular accumulation of PON activity in the presence of apoA-I WT was 0.095 ( 0.013 unit/mg of cell protein (n ) 7) compared to 0.034 ( 0.010 unit/mg of cell protein in the absence of apoA-I (n ) 7), a 2.79-fold increase in activity when apoA-I was incubated with the cells. Lipid-free apoA-I did not increase PON activity, while preformed nascent HDL increased PON activity only 30%, suggesting that maximal PON activity is lipid-dependent and requires coassembly of PON and apoA-I on nascent HDL. The cysteine mutations R10C, R27C, and R61C significantly increased (p < 0.01) PON activity 32.6% ( 14.7%, 31.6% ( 18.9%, and 27.4% ( 20%, respectively, over that of wild type (WT). No changes in PON activity were observed with apoA-I cysteine substitution mutations in the C-terminal portion of the protein.The data suggest that, for optimal PON activity, coassembly of the enzyme onto nascent HDL is required and that the N-terminal region of apoA-I may be important in the assembly process.

show abstract

“…A second chemical method uses SDS-PAGE to separate the apoB-100 and absorbance measurements of the stained bands (34). Immunoassay has also been used for quantitation of serum apoB-100 levels (35).…”

Section: Advantages Of a Ce-based Methods For Apob-100 Quantitationmentioning

confidence: 99%

Characterization and quantitation of apolipoprotein B-100 by capillary electrophoresis

Cruzado

Cockrill

McNeal

et al. 1998

Journal of Lipid Research

View full text Add to dashboard Cite

The interaction of low density lipoproteins (LDL) with different surfactants was studied by capillary electrophoresis (CE) and sucrose density gradient ultracentrifugation as part of developing a method for quantitation of apoB-100 in serum. A mixture of surfactants consisting of 70% sodium dodecyl sulfate (SDS), 25% sodium myristyl sulfate, and 5% sodium cetyl sulfate was found to delipidate LDL particles more effectively than pure SDS or sodium decyl sulfate. The delipidation products of LDL [apolipoprotein B-100 (apoB-100) and lipids] were resolved as two distinct peaks by CE when using a 3.5 m M 70% SDS mixture, 20% (v/v) acetonitrile, 50 m M sodium borate, pH 9.1 buffer. This CE method was also used to characterize apoB-100 derived from samples of lipoprotein [a] and very low density lipoproteins (VLDL). A CE-based quantitation method for apoB-100 was developed utilizing the observed linear relationship between apoB-100 concentration and its corrected 214 nm absorbance peak area measured on-line by CE. Concentration values of apoB-100 in LDL and VLDL samples were determined by CE and found to be accurate when compared to values obtained by immunoturbidimetric analysis and the Lowry method. Capillary electrophoresis can be used as a precise, accurate, and specific on-line method for the qualitative and quantitative analysis of the apoB-100 component of VLDL and LDL-related lipoproteins. -Cruzado, I.

show abstract

[31] Immunochemical methods for studying lipoprotein structure

Cited by 9 publications

References 78 publications

A Thyroid Hormone Binding Motif Is Evolutionarily Conserved in Apolipoproteins

A Thyroid Hormone Binding Motif Is Evolutionarily Conserved in Apolipoproteins

Cysteine Substitutions in Apolipoprotein A-I Primary Structure Modulate Paraoxonase Activity

Characterization and quantitation of apolipoprotein B-100 by capillary electrophoresis

Contact Info

Product

Resources

About