Effective encapsulation of proteins into size-controlled phospholipid vesicles using freeze-thawing and extrusion
Effective encapsulation of proteins into size-controlled phospholipid vesicles using freeze-thawing and extrusion
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DOI：
10.1021/bp0201004
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发表时间：
2003-09-01
影响因子：
2.9
通讯作者：
Tsuchida, E
中科院分区：
文献类型：
--
作者：
Sou, K;Naito, Y;Tsuchida, E
我们的目标是提高蛋白质的封装效率，在一个大小调节磷脂囊泡采用挤压法。混合脂质（1,2-二棕榈酰-sn-甘油-3-磷脂酰胆碱（DPPC））、胆固醇、1,5-二棕榈酰- l-谷氨酸- n-琥珀酸（DPEA）和1,2-二硬脂酰-sn-甘油-3-磷酸乙醇胺- n-[单甲氧基聚乙二醇（5000）](PEG-DSPE)，摩尔比分别为5,5,1和0.033，用NaOH溶液（7.6 mM）水化，得到多分散的多层囊泡分散体（50 nm至30 nm直径）。在脂质浓度为2 g dL（-1）和冷却速度为-140℃min（-1）的条件下，将多分散囊泡转化为平均直径约500 nm、尺寸分布相对狭窄的小囊泡。将冻融后囊泡的冻干粉再水化成浓缩蛋白溶液（羰基血红蛋白溶液，40 g dL(-1)），保留原囊泡的大小和大小分布。所产生的囊泡分散体在挤压过程中顺利地通过膜过滤器渗透。冻融囊泡的平均渗透速率比简单水化囊泡快30倍左右。在挤压过程中，蛋白质被包裹在直径为250 ~ 20 nm的重构囊泡中。
We are aiming to improve the encapsulation efficiency of proteins in a size-regulated phospholipid vesicle using an extrusion method. Mixed lipids (1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC), cholesterol, 1,5-dipalmitoyl-L-glutamate-N-succinic acid (DPEA), and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[monomethoxy poly(ethylene glycol) (5,000)] (PEG-DSPE) at a molar ratio of 5, 5, 1, and 0.033 were hydrated with a NaOH solution (7.6 mM) to obtain a polydispersed multilamellar vesicle dispersion (50 nm to 30 mum diameter). The polydispersed vesicles were converted to smaller vesicles having an average diameter of ca. 500 nm with a relatively narrow size distribution by freeze-thawing at a lipid concentration of 2 g dL(-1) and cooling rate of -140 degreesC min(-1). The lyophilized powder of the freeze-thawed vesicles was rehydrated into a concentrated protein solution (carbonyl hemoglobin solution, 40 g dL(-1)) and retained the size and size distribution of the original vesicles. The resulting vesicle dispersion smoothly permeated through the membrane filters during extrusion. The average permeation rate of the freeze-thawed vesicles was ca. 30 times faster than that of simple hydrated vesicles. During the extrusion process, proteins were encapsulated into the reconstructed vesicles with a diameter of 250 20 nm.