Multi a

Size and morphology of the particles were analyzed via transmission electron microscopy (TEM) studies using a ZEISS Leo912 microscope operated at an acceleration voltage of 120 kV. For sample preparation, diluted dispersions of the samples were prepared in ethanol, dropped onto a carbon-covered standard TEM grid (QUANTIFOIL Multi A) and dried under air. Phase purity and crystallinity of the samples was measured using a powder STOE-STADI MP X-Ray diffractometer (XRD) with Mo radiation (λ = 0.7093 Å) and measured peak patterns were compared to reference JCPDS files. Infrared spectra were acquired using a Perkin Elmer Fourier transform infrared (FTIR) spectrometer between 400 and 4000 cm⁻¹. UV-vis spectroscopy (LAMBDA 950 Perkin Elmer) was employed to detect the dye-labeled miRNA on the particle surface. Zeta potential and hydrodynamic diameter of the particles were measured using aqueous dispersions (distilled water with a pH of 6.5) of the particles employing a Malvern Zetasizer NanoZS.

Two batches of EM grids were prepared. The first batch used for collection of dataset 1 contained the in vivo pulled-out Ebp1–ribosome complex without addition of recombinant Ebp1. The second batch used for collection of dataset 2 contained the same purified ribosomes, but supplemented with recombinant Ebp1 (p48 isoform) to increase Ebp1 occupancy on the ribosomes. Right before freezing, Quantifoil Multi A holey carbon supported grids (Quantifoil, Multi A, 400 mesh) were glow-discharged for 10 s in oxygen atmosphere using a Solarus plasma cleaner (Gatan, Inc.). In total, 3 μL of freshly prepared samples (100 nM ribosomes without/with a eightfold excess of recombinant Ebp1) were directly applied to glow-discharged grids. Under a blot force of 0 at 100% humidity, the grids were blotted for 3 s with Whatman #1 filter papers using a Vitrobot Mark IV (FEI Company) operated at room temperature, and then immediately plunge-frozen in liquid ethane cooled with liquid nitrogen.

The crystal suspension was dispersed on a Cu holy carbon-coated TEM grid (MultiA, Quantifoil), and was left in vacuum until completely dehydrated.