Working Paper Short Note Version 1 This version is not peer-reviewed

Amino Acid Identification via Trna Charging, Deacylation, and Current Blockade in a Nanopore

Version 1 : Received: 14 August 2020 / Approved: 18 August 2020 / Online: 18 August 2020 (05:03:02 CEST)

How to cite: Sampath, G. Amino Acid Identification via Trna Charging, Deacylation, and Current Blockade in a Nanopore. Preprints 2020, 2020080374 Sampath, G. Amino Acid Identification via Trna Charging, Deacylation, and Current Blockade in a Nanopore. Preprints 2020, 2020080374

Abstract

A label-free four-step procedure to identify amino acids (AAs) is described. In Step 1 molecules of AA, a tRNA, the related tRNA synthetase (AARS), and ATP are confined in a cavity to enable charging of tRNA. In Step 2 the tRNA, charged or uncharged, is separated from the other reactants (ATP, AARS, and possibly AMP and free AAs). In Step 3 the separated tRNA is subjected to non-enzymatic deacylation to dissociate the AA if tRNA is charged. In Step 4 the products are transferred to an electrolytic cell with a nanopore, where a current blockade occurs if and only if there is a dissociated AA. If a blockade is observed AA is immediately known, identification is unambiguous because of tRNA superspecificity. The exact blockade size need not be known (for any AA) in Step 4 so there is no need to distinguish among different AAs. This is unlike other nanopore-based methods, which are crucially dependent on precise blockade level measurements. The procedure is done in parallel with N (20 ≤ N ≤ 61) copies of AA, N reservoir-cavities, and N pairs of e-cells each with a different tRNA, related AARS, and ATP. At least one of the tRNAs gets charged. Assuming no charging errors, if N1 tRNAs get charged in Step 1 and at least one of N2 (≤ N1) charged tRNAs is deacylated in Step 3, then with N1N2 tRNA molecules entering Step 1 AA can be identified with probability 1.

Subject Areas

AARS; deacylation; label-free; nanopore; current blockade; protein sequencing

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