The transport of bio-macromolecules across cell membranes, particularly the translocation of charged polymers like DNA and RNA through protein nanopores, is a crucial process. Single-molecule techniques have enabled the investigation of individual polymer translocation through biological and synthetic nanopores. This article focuses on the effect of macromolecular crowders, specifically polyethylene glycol (PEG), on the capture and translocation of DNA through α-hemolysin (αHL) protein nanopores. The presence of PEG crowders in the solution has been observed to enhance DNA capture and slow down translocation. A discrete-state stochastic model is proposed to describe the capture and translocation processes, taking into account the cooperative entry of PEG molecules into the nanopore and the electrostatic interactions between DNA and PEG crowders.
Facts
- The transport of bio-macromolecules across membranes is a vital process in cells.
- DNA and RNA translocation through protein nanopores is difficult to monitor in vivo.
- Single-molecule techniques allow the probing of individual polymer translocation through nanopores.
- Nanopores can act as sensors by detecting the charge distribution on nucleotides.
- Macromolecular crowders, such as PEG, can enhance DNA capture and slow down translocation through nanopores.
- PEG crowders can interact with biological molecules and affect their stability.
- The cooperative entry of PEG molecules within the nanopore is driven by cation-induced conformational changes.
The transport of bio-macromolecules across different membranes in cells is a vital life process. This includes the translocation of charged polymers, such as DNA and RNA, through protein nanopores. Single-molecule detection of DNA or RNA translocation through nanopores has been achieved by applying an electric field. The structure and conditions of nanopores can be modified to improve their efficiency and sensing abilities. One challenge is to increase the capture rates of polymers within the nanopore without accelerating their translocation, as faster translocation makes sequencing difficult.
Macromolecular crowders, such as poly(ethylene glycol) (PEG), can create a crowded environment that mimics cellular conditions. In this study, the impact of PEG crowders on DNA capture and translocation through nanopores is investigated. High concentrations of large-sized PEG crowders have been shown to enhance the capture of polypeptides and DNA within nanopores. The presence of PEG crowders can increase the sensitivity and detection of nucleic acids and proteins in solid-state nanopores.
A minimal discrete-state stochastic model is developed to describe the capture and translocation of a single DNA molecule within an α-hemolysin (αHL) protein nanopore in the presence of PEG crowders. The model considers cooperative entry of PEG molecules, electrostatic interactions between PEGs and DNA, and the impact of PEG crowders on capture and translocation rates. The theoretical predictions of the model are in good agreement with experimental observations.
The presence of PEG crowders within the nanopore affects the capture and translocation of DNA molecules. Cooperative entry of PEG molecules occurs, facilitated by cation-induced conformational changes in the nanopore. Electrostatic interactions between PEGs and DNA enhance capture rates and slow down translocation. The molecular-level understanding of how PEG crowding impacts polymer capture and translocation rates is important for accurate sequencing and the design of nanopore-based sensors.
- Bhawakshi Punia and Srabanti Chaudhury*
- SUBJECTS: Genetics, Kinetic parameters, Mathematical methods, Molecules, NanoporesShow Less
- ACS- The Journal of Physical Chemistry B
- Publication Date: June 9, 2023
