Formation of HDL-like complexes from apolipoprotein A-I(M) and DMPC.
Conditions for the preparation of reconstituted high density lipoproteins (HDLs) by incubation of the synthetic lipid dimyristoylphosphatidylcholine (DMPC) and recombinant apolipoprotein A-I(M) have been investigated as a function of ratio of incubation lipid to protein, incubation temperature and the lipid form (multilamellar (MLV) or small unilamellar (SUV) vesicles). The size distributions of the resultant lipid-protein complex particles from various incubations have been evaluated by native gel electrophoresis. Structural changes of the protein after incorporation into these complex particles have been estimated by CD. Thermal characteristics of the particles has been examined by DSC and correlated with CD results. Titration calorimetry has been used to obtain interaction parameters based on a simplified binding model. It is hypothesized that the major enthalpic step in the production of rHDLs is the primary association step between protein and lipid vesicles. It has been shown that by raising the temperature and incubation ratio, the formation of rHDL particles can be directed towards smaller size and a narrower size distribution. The results have been described on the basis of a model where formation of discoidal particles requires prior saturation of vesicle surface area by adsorbed protein, thus explaining differences between particles formed from MLVs and SUVs.
Suurkuusk
M
M
Department of Pharmaceutical Technology, Pharmacia & Upjohn AB, SP 19-5, S-112 87, Stockholm, Sweden. malin.suurkuusk@thermometric.se
Singh
S K
SK
D016632:Apolipoprotein A-I / Q000737:chemistry*; D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D016018:Least-Squares Analysis; D008075:Lipoproteins, HDL / Q000138:chemical synthesis* / Q000737:chemistry; D010316:Particle Size; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature; D017186:Titrimetry
D016632:Apolipoprotein A-I; D008075:Lipoproteins, HDL; D011994:Recombinant Proteins; C080631:apolipoprotein A-I Milano; D004134:Dimyristoylphosphatidylcholine