Super-resolution force spectroscopy reveals ribosomal motion at sub-nucleotide steps

Abstract

Probing biomolecular motion beyond a single nucleotide is technically challenging but fundamentally significant. We have developed super-resolution force spectroscopy (SURFS) with 0.5 pN force resolution and revealed that the ribosome moves by half a nucleotide upon the formation of the pre-translocation complex, which is beyond the resolution of other techniques.

High‐Resolution DNA Dual‐Rulers Reveal a New Intermediate State in Ribosomal Frameshifting

“…Meanwhile on the technical front, we invented super‐resolution force spectroscopy (SURFS) by implementing acoustic radiation force, which was automatable and more precise than the centrifugal force used in FIRMS [14] . With 0.5 pN force resolution, we observed subnucleotide conformational difference between two different ribosome states within one translocation step [15] . Compared to other force‐based techniques for ribosome research, mainly atomic force microscopy (AFM) and optical tweezers, [16,17] our approaches have the unique advantages of resolving different reading frames within the same sample, and probing both the moving edges of the ribosome.…”

Noncovalent force spectroscopy using wide-field optical and diamond-based magnetic imaging

“…FIRMS offers high force resolution that can precisely distinguish noncovalent intermolecular interactions, for example those of DNA duplexes with a single basepair difference. 6,7 However, FIRMS currently uses vapor-based atomic magnetometers for detection, with sensor sizes on the order of millimeters. Due to the size mismatch, the atomic magnetometers cannot detect individual magnetic beads that are used to label the biomolecules.…”

Probing drug-DNA interactions using super-resolution force spectroscopy

“…[13][14][15] Recently, we showed super-resolution force spectroscopy (SURFS) that uses acoustic radiation force to precisely distinguish different intermolecular bonds, consequently revealing valuable mechanistic information of complex biological processes. 16,17 Here, we report a unique application of SURFS in biophysics, as well as the critical technical aspects during the integration of an automated magnetometer and an ultrasound generator. The binding of three small ligands onto the minor grooves of two different DNA duplexes was precisely determined, with an order of magnitude better resolution than other force spectroscopic techniques.…”

“…The experimental procedures were similar to previous studies and are provided in the supplementary material. 17,20 Figure 3(a) shows the relative magnetic signal vs. acoustic radiation force for a 11-basepair (bp) DNA containing a AAAAA segment for the three ligands. The DNA sequence was 5 0 -GCGAAAAACGC.…”

“…The force values were obtained according to a previous calibration using a series of DNA duplexes, in which the same molecular systems were dissociated by both ultrasound and centrifugal force. 17 The calibration yielded the empirical formula of F ¼ 736 Â…”