Topology of mRNA chain in isolated eukaryotic double-row polyribosomes
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In the process of protein synthesis, the translating ribosomes of eukaryotic cells form polyribosomes that are found to be multiplex functional complexes possessing elements of ordered spatial organization. As revealed by a number of electron microscopy studies, the predominant visible configurations of the eukaryotic polyribosomes are circles (circular polyribosomes) and two-stranded formations (so-called double-row polyribosomes). The “long” (i.e. heavy loaded) polyribosomes are usually represented by double-row structures, which can be interpreted as either topologically circular (“col-lapsed rings”), or topologically linear (zigzags or helices). In the present work we have analyzed the mRNA path within the eukaryotic polyribosomes, isolated from a wheat germ cell-free translation system, by integrating two approaches: the visualization of mRNA ends in polyribosomes by marking them with gold nanoparticles (3′-end) and initiating 40S subunits (5′-end), as well as by the cryoelectron tomography. Examination of the location of the mRNA markers in polyribosomes and mutual orientation of ribosomes in them has shown that the double-row polyribosomes of the same sample can have both circular and linear arrangements of their mRNA.
Key wordseukaryotic polyribosomes mRNA circular translation cryoelectron tomography
continuous exchange cell-free system
eukaryotic initiation factor
- “MAFITC 10 nm gold”
anti-fluorescein isothiocyanate antibody conjugated with 10 nm gold particles
untranslated region of mRNA
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