Encapsulation of a biological molecule or a molecular complex in a vesicle provides a means of biofriendly immobilization for single molecule studies and further enables new types of analysis if the vesicles are permeable. We previously reported on using DMPC (dimyristoylphosphatidylcholine) vesicles for realizing porous bioreactors. Here, we describe a different strategy for making porous vesicles using a bacterial pore-forming toxin, α-hemolysin. Using RNA folding as a test case, we demonstrate that protein-based pores can allow exchange of magnesium ions through the vesicle wall while keeping the RNA molecule inside. Flow measurements indicate that the encapsulated RNA molecules rapidly respond to the change in the outside buffer condition. The approach was further tested by coencapsulating a helicase protein and its single-stranded DNA track. The DNA translocation activity of E. coli Rep helicase inside vesicles was fueled by ATP provided outside the vesicle, and a dramatically higher number of translocation cycles could be observed due to the minuscule vesicle volume that facilitates rapid rebinding after dissociation. These pores are known to be stable over a wide range of experimental conditions, especially at various temperatures, which is not possible with the previous method using DMPC vesicles. Moreover, engineered mutants of the utilized toxin can potentially be exploited in the future applications.
proliferation in smaller islets than in larger islets. In adulthood, however, every beta cell in an islet of arbitrary size ultimately has an equally small proliferation potential. In addition to this limited islet growth, fission of large islets occurred most actively at postnatal week 3, and contributed to maintaining a limited range of islet sizes. On the other hand, in a tumor (insulinoma) progression model, we found unlimited islet growth, with especially accelerated cell proliferation in larger islets. We conclude that islet size is constrained by preferential growth of small islets and fission of large islets in the early postnatal period, and a low rate of proliferation in maturity.
The geographic coordinate system on the sphere S 2 . . . . . . . 8.2 Generation of a singularity free grid on S 2 . . . . . . . . . . . . 8.3 The grid on the net of the three-and four-dimensional cubes. .
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