Probing the conformation of a human apolipoprotein C-1 by amino acid substitutions and trimethylamine-N-oxide.


Abstract

To test, at the level of individual amino acids, the conformation of an exchangeable apolipoprotein in aqueous solution and in the presence of an osmolyte trimethylamine-N-oxide (TMAO), six synthetic peptide analogues of human apolipoprotein C-1 (apoC-1, 57 residues) containing point mutations in the predicted alpha-helical regions were analyzed by circular dichroism (CD). The CD spectra and the melting curves of the monomeric wild-type and plasma apoC-1 in neutral low-salt solutions superimpose, indicating 31 +/- 4% alpha-helical structure at 22 degrees C that melts reversibly with T(m,WT) = 50 +/- 2 degrees C and van't Hoff enthalpy deltaH(v,WT)(Tm) = 18 +/- 2 kcal/mol. G15A substitution leads to an increased alpha-helical content of 42 +/- 4% and an increased T(m,G15A) = 57 +/- 2 degrees C, which corresponds to stabilization by delta deltaG(app) = +0.4 +/- 1.5 kcal/mol. G15P mutant has approximately 20% alpha-helical content at 22 degrees C and unfolds with low cooperativity upon heating to 90 degrees C. R23P and T45P mutants are fully unfolded at 0-90 degrees C. In contrast, Q31P mutation leads to no destabilization or unfolding. Consequently, the R23 and T45 locations are essential for the stability of the cooperative alpha-helical unit in apoC-1 monomer, G15 is peripheral to it, and Q31 is located in a nonhelical linker region. Our results suggest that Pro mutagenesis coupled with CD provides a tool for assigning the secondary structure to protein groups, which should be useful for other self-associating proteins that are not amenable to NMR structural analysis in aqueous solution. TMAO induces a reversible cooperative coil-to-helix transition in apoC-1, with the maximal alpha-helical content reaching 74%. Comparison with the maximal alpha-helical content of 73% observed in lipid-bound apoC-1 suggests that the TMAO-stabilized secondary structure resembles the functional lipid-bound apolipoprotein conformation. Study holds ProTherm entries: 6818, 6819 Extra Details: amphipathic alpha-helix; CD spectroscopy;,plasma lipoproteins; protein folding; osmolyte

Submission Details

ID: DmoqsPun3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:32 p.m.

Version: 1

Publication Details
Gursky O,Protein Sci. (1999) Probing the conformation of a human apolipoprotein C-1 by amino acid substitutions and trimethylamine-N-oxide. PMID:10548051
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1ALF 1995-04-20 CONFORMATION OF TWO PEPTIDES CORRESPONDING TO HUMAN APOLIPOPROTEIN C-I RESIDUES 7-24 AND 35-53 IN THE PRESENCE OF SODIUM DODECYLSULFATE BY CD AND NMR SPECTROSCOPY
1ALE 1995-04-20 CONFORMATION OF TWO PEPTIDES CORRESPONDING TO HUMAN APOLIPOPROTEIN C-I RESIDUES 7-24 AND 35-53 IN THE PRESENCE OF SODIUM DODECYLSULFATE BY CD AND NMR SPECTROSCOPY
1OPP 1998-05-13 PEPTIDE OF HUMAN APOLIPOPROTEIN C-I RESIDUES 1-38, NMR, 28 STRUCTURES
1JFN 2002-06-28 SOLUTION STRUCTURE OF HUMAN APOLIPOPROTEIN(A) KRINGLE IV TYPE 6
2FEB 2006-12-26 NMR Solution Structure, Dynamics and Binding Properties of the Kringle IV Type 8 module of apolipoprotein(a)
1IOJ 1998-08-12 HUMAN APOLIPOPROTEIN C-I, NMR, 18 STRUCTURES
1I71 2001-06-13 1.45 HIGH RESOLUTION CRYSTAL STRUCTURE OF APOLIPOPROTEIN(A) KRINGLE IV TYPE 7: INSIGHTS INTO LIGAND BINDING
4BVC 2014-07-09 1.6 Identification of small molecule inhibitors selective for apo(a) kringles KIV-7, KIV-10 and KV.
4BVD 2014-07-16 1.68 Identification of small molecule inhibitors selective for apo(a) kringles KIV-7, KIV-10 and KV.
4BV7 2014-07-09 1.7 Identification of small molecule inhibitors selective for apo(a) kringles KIV-7, KIV-10 and KV.
3KIV 1999-05-18 1.8 RECOMBINANT KRINGLE IV-10/M66 VARIANT OF HUMAN APOLIPOPROTEIN(A)
4BVV 2014-07-16 1.8 Identification of small molecule inhibitors selective for apo(a) kringles KIV-7, KIV-10 and KV.
6DVU 2018-12-26 1.8 Structure of the Monoclinic-1 (Monocl-1) Crystal Form of Human Apolipoprotein C1
6DXR 2018-12-26 2.0 Structure of the Monoclinic-2 (Monocl-2) Crystal Form of Human Apolipoprotein C1
4BVW 2014-07-16 2.0 Identification of small molecule inhibitors selective for apo(a) kringles KIV-7, KIV-10 and KV.
4BV5 2014-07-09 2.1 Identification of small molecule inhibitors selective for apo(a) kringles KIV-7, KIV-10 and KV.
1KIV 1999-05-18 2.1 RECOMBINANT KRINGLE IV-10/M66 VARIANT OF HUMAN APOLIPOPROTEIN(A)
4KIV 1999-05-18 2.2 RECOMBINANT KRINGLE IV-10/W72R MUTANT OF HUMAN APOLIPOPROTEIN(A)
6NF3 2018-12-26 2.33 Structure of the Monoclinic-3 (Monocln-3) Crystal Form of Human Apolipoprotein C1
6DZ6 2018-12-26 3.0 Structure of the Orthorhombic (Orthrhmb) Crystal Form of Human Apolipoprotein C1

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
92.1 Apolipoprotein C-I P0CE40 APO1B_PONAB
98.2 Apolipoprotein C-I P0CE38 APO1B_PANTR
100.0 Apolipoprotein C-I P02654 APOC1_HUMAN
100.0 Apolipoprotein(a) P08519 APOA_HUMAN