Charge transport in individual short base stacked single-stranded RNA molecules.

Charge transport in biomolecules is crucial for many biological and technological applications, including biomolecular electronics devices and biosensors. RNA has become the focus of research because of its importance in biomedicine, but its charge transport properties are not well understood. Here, we use the Scanning Tunneling Microscopy-assisted molecular break junction method to measure the electrical conductance of particular 5-base and 10-base single-stranded (ss) RNA sequences capable of base stacking. These ssRNA sequences show single-molecule conductance values around [Formula: see text] ([Formula: see text]), while equivalent-length ssDNAs result in featureless conductance histograms. Circular dichroism (CD) spectra and MD simulations reveal the existence of extended ssRNA conformations versus folded ssDNA conformations, consistent with their different electrical behaviors. Computational molecular modeling and Machine Learning-assisted interpretation of CD data helped us to disentangle the structural and electronic factors underlying CT, thus explaining the observed electrical behavior differences. RNA with a measurable conductance corresponds to sequences with overall extended base-stacking stabilized conformations characterized by lower HOMO energy levels delocalized over a base-stacking mediating CT pathway. In contrast, DNA and a control RNA sequence without significant base-stacking tend to form closed structures and thus are incapable of efficient CT.

A single-molecule RNA electrical biosensor for COVID-19.

The COVID-19 pandemic shows a critical need for rapid, inexpensive, and ultrasensitive early detection methods based on biomarker analysis to reduce mortality rates by containing the spread of epidemics. This can be achieved through the electrical detection of nucleic acids at the single-molecule level. In particular, the scanning tunneling microscopic-assisted break junction (STM-BJ) method can be utilized to detect individual nucleic acid molecules with high specificity and sensitivity in liquid samples. Here, we demonstrate single-molecule electrical detection of RNA coronavirus biomarkers, including those of SARS-CoV-2 as well as those of different variants and subvariants. Our target sequences include a conserved sequence in the human coronavirus family, a conserved target specific for the SARS-CoV-2 family, and specific targets at the variant and subvariant levels. Our results demonstrate that it is possible to distinguish between different variants of the COVID-19 virus using electrical conductance signals, as recently suggested by theoretical approaches. Our results pave the way for future miniaturized single-molecule electrical biosensors that could be game changers for infectious diseases and other public health applications.

Electrical detection of RNA cancer biomarkers at the single-molecule level.

Cancer is a significant healthcare issue, and early screening methods based on biomarker analysis in liquid biopsies are promising avenues to reduce mortality rates. Electrical detection of nucleic acids at the single molecule level could enable these applications. We examine the electrical detection of RNA cancer biomarkers (KRAS mutants G12C and G12V) as a single-molecule proof-of-concept electrical biosensor for cancer screening applications. We show that the electrical conductance is highly sensitive to the sequence, allowing discrimination of the mutants from a wild-type KRAS sequence differing in just one base. In addition to this high specificity, our results also show that these biosensors are sensitive down to an individual molecule with a high signal-to-noise ratio. These results pave the way for future miniaturized single-molecule electrical biosensors that could be groundbreaking for cancer screening and other applications.

Single-molecule conductance of double-stranded RNA oligonucleotides.

RNA oligonucleotides are crucial for a range of biological functions and in many biotechnological applications. Herein, we measured, for the first time, the conductance of individual double-stranded (ds)RNA molecules and compared it with the conductance of single DNA : RNA hybrids. The average conductance values are similar for both biomolecules, but the distribution of conductance values shows an order of magnitude higher variability for dsRNA, indicating higher molecular flexibility of dsRNA. Microsecond Molecular Dynamics simulations explain this difference and provide structural insights into the higher stability of DNA : RNA duplex with atomic level of detail. The rotations of 2'-OH groups of the ribose rings and the bases in RNA strands destabilize the duplex structure by weakening base stacking interactions, affecting charge transport, and making single-molecule conductance of dsRNA more variable (dynamic disorder). The results demonstrate that a powerful combination of state-of-the-art biomolecular electronics techniques and computational approaches can provide valuable insights into biomolecules' biophysics with unprecedented spatial resolution.

RNA BioMolecular Electronics: towards new tools for biophysics and biomedicine.

The last half-century has witnessed the birth and development of a new multidisciplinary field at the edge between materials science, nanoscience, engineering, and chemistry known as Molecular Electronics. This field deals with the electronic properties of individual molecules and their integration as active components in electronic circuits and has also been applied to biomolecules, leading to BioMolecular Electronics and opening new perspectives for single-molecule biophysics and biomedicine. Herein, we provide a brief introduction and overview of the BioMolecular electronics field, focusing on nucleic acids and potential applications for these measurements. In particular, we review the recent demonstration of the first single-molecule electrical detection of a biologically-relevant nucleic acid. We also show how this could be used to study biomolecular interactions and applications in liquid biopsy for early cancer detection, among others. Finally, we discuss future perspectives and challenges in the applications of this fascinating research field.

Unraveling the Excited-State Dynamics and Light-Harvesting Functions of Xanthophylls in Light-Harvesting Complex II Using Femtosecond Stimulated Raman Spectroscopy.

Photosynthesis in plants starts with the capture of photons by light-harvesting complexes (LHCs). Structural biology and spectroscopy approaches have led to a map of the architecture and energy transfer pathways between LHC pigments. Still, controversies remain regarding the role of specific carotenoids in light-harvesting and photoprotection, obligating the need for high-resolution techniques capable of identifying excited-state signatures and molecular identities of the various pigments in photosynthetic systems. Here we demonstrate the successful application of femtosecond stimulated Raman spectroscopy (FSRS) to a multichromophoric biological complex, trimers of LHCII. We demonstrate the application of global and target analysis (GTA) to FSRS data and utilize it to quantify excitation migration in LHCII trimers. This powerful combination of techniques allows us to obtain valuable insights into structural, electronic, and dynamic information from the carotenoids of LHCII trimers. We report spectral and dynamical information on ground- and excited-state vibrational modes of the different pigments, resolving the vibrational relaxation of the carotenoids and the pathways of energy transfer to chlorophylls. The lifetimes and spectral characteristics obtained for the S1 state confirm that lutein 2 has a distorted conformation in LHCII and that the lutein 2 S1 state does not transfer to chlorophylls, while lutein 1 is the only carotenoid whose S1 state plays a significant energy-harvesting role. No appreciable energy transfer takes place from lutein 1 to lutein 2, contradicting recent proposals regarding the functions of the various carotenoids (Son et al. Chem. 2019, 5 (3), 575-584). Also, our results demonstrate that FSRS can be used in combination with GTA to simultaneously study the electronic and vibrational landscapes in LHCs and pave the way for in-depth studies of photoprotective conformations in photosynthetic systems.