title: MspA nanopore as a single-molecule tool: From sequencing to SPRNT.
abstract: Single-molecule picometer resolution nanopore tweezers (SPRNT) is a new tool for analyzing the motion of nucleic acids through molecular motors. With SPRNT, individual enzymatic motions along DNA as small as 40pm can be resolved on sub-millisecond time scales. Additionally, SPRNT reveals an enzyme's exact location with respect to a DNA strand's nucleotide sequence, enabling identification of sequence-specific behaviors. SPRNT is enabled by a mutant version of the biological nanopore formed by Mycobacterium smegmatis porin A (MspA). SPRNT is strongly rooted in nanopore sequencing and therefore requires a solid understanding of basic principles of nanopore sequencing. Furthermore, SPRNT shares tools developed for nanopore sequencing and extends them to analysis of single-molecule kinetics. As such, this review begins with a brief history of our work developing the nanopore MspA for nanopore sequencing. We then describe the underlying principles of SPRNT, how it works in detail, and propose some potential future uses. We close with a comparison of SPRNT to other techniques and we present the methods that will enable others to use SPRNT.
authors.affiliation: University of Washington, Department of Physics, 3910 15th Ave NE, Seattle, WA 98195, USA.
authors.forename: Andrew H
authors.initials: AH
authors.lastname: Laszlo
authors.affiliation: University of Washington, Department of Physics, 3910 15th Ave NE, Seattle, WA 98195, USA.
authors.forename: Ian M
authors.initials: IM
authors.lastname: Derrington
authors.affiliation: University of Washington, Department of Physics, 3910 15th Ave NE, Seattle, WA 98195, USA. Electronic address: jens@phys.washington.edu.
authors.forename: Jens H
authors.initials: JH
authors.lastname: Gundlach
mesh_terms: D007700:Kinetics; D020102:Mycobacterium smegmatis / Q000201:enzymology; D058608:Nanopores / Q000648:ultrastructure; D009696:Nucleic Acids / Q000737:chemistry* / Q000235:genetics; D052898:Optical Tweezers; D018272:Porins / Q000737:chemistry* / Q000235:genetics; D000072760:Single Molecule Imaging / Q000379:methods*
chemical_list: D009696:Nucleic Acids; D018272:Porins; C121838:mspA protein, Mycobacterium smegmatis
keywords: Force spectroscopy; Helicase; Kinetics; Polymerase; Single-molecule