Preparation and characterization of reactive and stable glucose oxidase-containing liposomes modulated with detergent.
Preparation and characterization of reactive and stable glucose oxidase-containing liposomes modulated with detergent.
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用去垢剂调节的含有反应性和稳定的葡萄糖氧化酶的脂质体的制备和表征。
DOI：
10.1002/bit.10514
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发表时间：
2003
影响因子：
3.8
通讯作者：
K. Nakao
中科院分区：
文献类型：
--
作者：
M. Yoshimoto;Shaoqing Wang;K. Fukunaga;P. Walde;R. Kuboi;K. Nakao
制备并表征了含葡萄糖氧化酶的脂质体（GOL）以及洗涤剂调节的含葡萄糖氧化酶的脂质体，不仅关注外部添加葡萄糖后脂质体的反应性，而且关注包埋的葡萄糖氧化酶（GO）从脂质体的泄漏，目的是开发反应性和稳定的脂质体GO系统。制备的GOL的膜由1-棕榈酰-2-油酰-sn-甘油-3-磷酸胆碱（POPC）组成，并用Triton X-100或胆酸盐调节。在不存在添加的去污剂的情况下，没有观察到GO从GOL泄漏，而其酶活性非常低（低葡萄糖渗透性）。作为去污剂调节的脂质体，以不同的有效去污剂/POPC摩尔比（R（e））制备混合POPC/Triton X-100和混合POPC/胆酸盐脂质体（分别缩写为TL和CL），所述有效去污剂/POPC摩尔比（R（e））范围为R（e）= 0至R（e）（sat）（R（e）（sat）是脂质体膜被去污剂饱和时的R（e）的临界值）。GO负载的TL（缩写为GOTL）或GO负载的CL（GOCL）的反应性随着脂质体中各自去污剂含量的增加而急剧增加。在GOTL的情况下，在R（e）（sat）= 0.40时，测量到高反应性，同时具有高程度的GO泄漏，表明观察到的酶促反应主要由泄漏的GO催化，这是由Triton X-100与POPC膜的相互作用引起的。另一方面，在R（e）（sat）= 0.43下制备的GOCL显示出相对高的反应性，仅具有小程度的GO泄漏，这表明大部分酶反应受到葡萄糖渗透穿过GOCL双层的限制。发现GOCL中的GO泄漏主要发生在GOCL制备过程中脂质体膜的重排期间（将GOL和胆酸盐混合）。膜相关的DPH（1，6-二苯基-1，3，5-己三烯）的荧光偏振测量表明，CL制备的修饰POPC与胆酸盐没有导致膜流动性的急剧变化，表明相互作用的胆酸盐分子没有深入渗透到POPC双层。总之，清楚地表明，GOL的膜渗透性可以通过将其与一定量的胆酸盐混合以形成具有最小酶泄漏的高反应性和稳定的GOCL而非常简单地调节。
Glucose oxidase-containing liposomes (GOL) as well as detergent-modulated glucose oxidase-containing liposomes were prepared and characterized, focusing not only on the reactivity of the liposomes upon external addition of glucose but also on the leakage of the entrapped glucose oxidase (GO) from the liposomes with the aim of developing a reactive and stable liposomal GO system. The membranes of the GOL prepared were composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and modulated with either Triton X-100 or cholate. In the absence of added detergent, no GO leakage from the GOL was observed while its enzymatic activity was very low (low glucose permeability). As detergent-modulated liposomes, mixed POPC/Triton X-100 and mixed POPC/cholate liposomes (abbreviated as TL and CL, respectively) were prepared at different effective detergent/POPC molar ratios (R(e)) ranging from R(e) = 0 to R(e) = R(e) (sat) (R(e) (sat) is the critical value of R(e) at which the liposome membrane is saturated with detergent). The reactivity of GO-loaded TL (abbreviated as GOTL) or GO-loaded CL (GOCL) increased drastically with increase in the respective detergent content in the liposomes. In the case of GOTL, at R(e) (sat) = 0.40, a high reactivity was measured with a simultaneous high extent of GO leakage, suggesting that the observed enzymatic reaction was catalyzed mainly by leaked GO, caused by the interaction of Triton X-100 with the POPC membrane. On the other hand, GOCL prepared at R(e) (sat) = 0.43 showed relatively high reactivity with only a small extent of GO leakage, suggesting that most of the enzyme reaction was limited by the glucose permeation across the bilayers of GOCL. The GO leakage from GOCL was found to occur mostly during the rearrangement of the liposomal membrane during the preparation of the GOCL (mixing the GOL and cholate). Fluorescence polarization measurements of membrane-associated DPH (1,6-diphenyl-1,3,5-hexatriene) indicated that CL prepared by modifying POPC with cholate did not lead to a drastic change in membrane fluidity, indicating that the interacting cholate molecules did not penetrate deeply into the POPC bilayers. In summary, it was clearly shown that the membrane permeability of GOL can be quite simply modulated by mixing it with a certain amount of cholate to form highly reactive and stable GOCL with minimal enzyme leakage.
DOI：
10.1016/0005-2736(91)90295-j
发表时间：
1991-01-30
期刊：
BIOCHIMICA ET BIOPHYSICA ACTA
影响因子：
--
作者：
MACDONALD, RC;MACDONALD, RI;HU, LR
通讯作者：
HU, LR