Size is a major determinant of dissociation and denaturation behaviour of reconstituted high-density lipoproteins.
Lipid-free apolipoprotein A-I (apoA-I) and A-I(Milano) (A-I(M)) were compared for their denaturation behaviour by running across transverse gradients of a chaotrope, urea, and of a ionic detergent, SDS. For both apo A-I and monomeric apoA-I(M) in the presence of increasing concentrations of urea the transition from high to low mobility had a sigmoidal course, whereas for dimeric A-I(M)/A-I(M) a non-sigmoidal shape was observed. The co-operativity of the unfolding process was lower for dimeric A-I(M)/A-I(M) than for apoA-I or for monomeric apoA-I(M). A slightly higher susceptibility to denaturation was observed for dimeric A-I(M)/A-I(M) than for monomeric apoA-I(M). A similar behaviour of A-I(M)/A-IM versus apoA-I(M) was observed in CD experiments. Large- (12.7/12.5 nm) and small- (7.8 nm) sized reconstituted high-density lipoproteins (rHDL) containing either apoA-I or A-I(M)/A-I(M) were compared with respect to their protein-lipid dissociation behaviour by subjecting them to electrophoresis in the presence of urea, of SDS and of a non-ionic detergent, Nonidet P40. A higher susceptibility to dissociation of small-sized versus large-sized rHDL, regardless of the apolipoprotein component, was observed in all three instances. Our data demonstrate that the differential plasticity of the various classes of rHDL is a function of their size; the higher stability of 12.5/12.7 nm rHDL is likely connected to the higher number of protein-lipid and lipid-lipid interactions in larger as compared with smaller rHDL.
Gianazza
Elisabetta
E
Gruppo di Studio per la Proteomica e la Struttura delle Proteine, Dipartimento di Scienze Farmacologiche, Università degli Studi di Milano, via G. Balzaretti 9, I-20133 Milano, Italy. elisabetta.gianazza@unimi.it
Eberini
Ivano
I
Sirtori
Cesare R
CR
Franceschini
Guido
G
Calabresi
Laura
L
D002942:Circular Dichroism; D003902:Detergents / Q000494:pharmacology; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D007477:Ions; D007700:Kinetics; D008075:Lipoproteins, HDL / Q000378:metabolism*; D010713:Phosphatidylcholines / Q000494:pharmacology; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry / Q000494:pharmacology; D014508:Urea / Q000494:pharmacology
D003902:Detergents; D007477:Ions; D008075:Lipoproteins, HDL; D010713:Phosphatidylcholines; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D014508:Urea