| Popis: |
Apoprotein B, the major apoprotein of normal human low density lipoprotein (LDL) was solubilized with sodium deoxycholate (NaDC). The protein was recombined with the phospholipid dimyristoyl phosphatidylcholine (DMPC) to produce a complex of DMPC-apoB (4:1 w/w). (Biochemistry. 22: 3170-3178. 1983). Carboxyfluorescein and [3H]dextran entrapment studies show the DMPC-apoB 4:1 (w/w) complex to encapsulate an aqueous volume of 0.17 microliter/mumol of DMPC. From the chemistry and morphology of the complex and the evidence that the complex possesses an encapsulated volume, the most appropriate structural model for this assembly is that of a phospholipid single bilayer vesicle into which apoB is incorporated. Differential scanning calorimetry (DSC) and circular dichroic spectroscopy (CD) were used to investigate the physical properties of apoB in the mixed micellar complex with NaDC and in the vesicular DMPC-apoB complex. CD studies of apoB in NaDC mixed micelles show that apoB exhibits a similar secondary structure as apoB of native LDL over the temperature range 5-30 degrees C. Reversible structural changes occur between 30 and 50 degrees C. However, above 50 degrees C, disruption of the micellar particle and endothermic protein unfolding and denaturation occur with a Tmax of 52 degrees C and an enthalpy of 0.22 cal/g apoB, as shown by DSC. The DMPC-apoB complex exhibits a reversible thermal transition centered at 24 degrees C (delta H = 3.34 Kcal/mol DMPC) which is associated with the order-disorder transition of the hydrocarbon chains of DMPC. An endothermic transition occurs over the range 53-70 degrees C (delta H = 2.09 cal/g apoB) which, as shown by CD and turbidity study, corresponds to protein unfolding-denaturation and particle disruption. CD shows that apoB in the vesicular environment undergoes a series of conformational changes. The major alterations occur over the temperature range of the order-disorder transition of the phospholipid. Between 37-60 degrees C, the conformation is similar to that observed in native LDL. |
