詳細介紹

Frankia are gram-positive, filamentous, nitrogen-fixing actinobacteria that are symbiotic with over 200

different species of plants. Frankia produce three cell types: vegetative hyphae, spores located in

sporangia, and the unique lipid-enveloped cellular structures, termed vesicles. Vesicles are formed

inside plant cell nodules, or in culture under nitrogen limiting conditions, and act as specialized structures

for the nitrogen fixation process. The mature vesicle is surrounded by an envelope that extends down the

stalk of the vesicle past the basal septum, which separates the vesicle from the hyphae. Techniques have

been developed for the isolation and urification of intact vesicles from Frankia grown in culture [1,2,3]. Initial

investigations on the properties of purified vesicles have focused on nitrogen metabolism [3,4]. Protein

extraction from vesicles has historically been difficult and a significant bottleneck to proteomic studies

because vesicles are resilient structures that are difficult to disrupt. Reliable and comprehensive proteomic

maps of Frankia hyphae and vesicles can only be assured when proteins are isolated reproducibly and in

a manner in which they are accuray represented in the downstream analysis. For example, 2DGE can be

an accurate representation of a proteome only if the entire protein constituency of cells is recovered during

the sample preparation process. While French press treatment effectively lyses hyphae, it fails to disrupt

Frankia vesicles. Pressure Cycling Technology （PCT） used in combination with a specialized lysis reagent

effectively disrupted purified vesicles enabling further investigation of the proteome of the vesicles.

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