Osawa Masaki, Erickson Harold P
Department of Cell Biology, Duke University Medical Center, Durham, North Carolina, USA.
Methods Enzymol. 2009;464:3-17. doi: 10.1016/S0076-6879(09)64001-5.
We have developed a system for producing tubular multilamellar liposomes that incorporate the protein FtsZ on the inside. We start with a mixture of spherical multilamellar liposomes with FtsZ initially on the outside. Shearing forces generated by applying a coverslip most likely distort some of the spherical liposomes into a tubular shape, and causes some to leak and incorporate FtsZ inside. We describe protocols for liposome preparation, and for preparing membrane-targeted FtsZ that can assemble contractile Z rings inside the tubular liposomes. We also describe the characterization of the multilamellar liposomes in terms of the permeability or leakiness for a small fluorescent dye and larger protein molecules. These liposomes may be useful for reconstitution of other biological systems.
我们开发了一种用于生产内部包含FtsZ蛋白的管状多层脂质体的系统。我们从最初FtsZ位于外部的球形多层脂质体混合物开始。通过施加盖玻片产生的剪切力很可能将一些球形脂质体扭曲成管状,并导致一些脂质体泄漏并使FtsZ进入内部。我们描述了脂质体制备的方案,以及制备可在管状脂质体内组装收缩性Z环的膜靶向FtsZ的方案。我们还描述了多层脂质体在对一种小荧光染料和较大蛋白质分子的渗透性或泄漏性方面的表征。这些脂质体可能对其他生物系统的重构有用。