Devika, Vikraman and Remya , Satheesan and Santhosh Kumar, K. and Mahendran, K R (2020) Nanopore Passport Control for Substrate-Specific Translocation. ACS nano, 14 (2). pp. 2285-2295. ISSN 1936-0851

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Abstract

Membrane protein pores have demonstrated applications in nanobiotechnology and single-molecule chemistry for effective detection of biomolecules. Here, we define the molecular basis of carbohydrate polymers translocation through a substrate-specific bacterial nanopore, CymA, which has a 15-residue N terminus segment inside the pore, restricting its diameter. Using single-channel recordings, we determined the kinetics of cationic cyclic oligosaccharide binding and elucidated the translocation mechanism across the pore in real-time. The cationic cyclic hexasaccharide binds to the densely packed negatively charged residues at the extracellular side of the pore with high affinity, facilitating its entry into the pore driven by the applied voltage. Further, the dissociation rate constant increased with increasing voltages, indicating unidirectional translocation toward the pore exit. Specifically, a larger cationic cyclic octasaccharide rapidly blocked the pore more effectively, resulting in the complete closure of the pore with increasing voltage, implying only strong binding. Further, we show that uncharged oligosaccharides exclusively bind to the extracellular side of the pore and the electroosmotic flow most likely drives their translocation. We propose that CymA favors selective translocation of cyclic hexasaccharide and linear maltooligosaccharides due to an asymmetrical charge pattern and the N terminus that regulates the substrate transport. We suggest that this substrate-specific nanopore with sophisticated geometry will be useful for complex biopolymer characterization.

Item Type:	Article
Uncontrolled Keywords:	nanopore membrane pores oligosaccharides translocation binding kinetics
Subjects:	Chemical Biology
Depositing User:	Central Library RGCB
Date Deposited:	24 Nov 2020 06:12
Last Modified:	24 Nov 2020 06:12
URI:	http://rgcb.sciencecentral.in/id/eprint/987

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