Direct electrochemical observation of glucosidase activity in isolated single lysosomes from a living cell

Rongrong Pan, Mingchen Xu, James D. Burgess, Dechen Jiang, Hong Yuan Chen

Research output: Contribution to journalArticle

33 Scopus citations

Abstract

The protein activity in individual intracellular compartments in single living cells must be analyzed to obtain an understanding of protein function at subcellular locations. The current methodology for probing activity is often not resolved to the level of an individual compartment, and the results provide an extent of reaction that is averaged from a group of compartments. To address this technological limitation, a single lysosome is sorted from a living cell via electrophoresis into a nanocapillary designed to electrochemically analyze internal solution. The activity of a protein specific to lysosomes, β-glucosidase, is determined by the electrochemical quantification of hydrogen peroxide generated from the reaction with its substrate and the associated enzymes preloaded in the nanocapillary. Sorting and assaying multiple lysosomes from the same cell shows the relative homogeneity of protein activity between different lysosomes, whereas the protein activity in single lysosomes from different cells of the same type is heterogeneous. Thus, this study for the analysis of protein activity within targeted cellular compartments allows direct study of protein function at subcellular resolution and provides unprecedented information about the homogeneity within the lysosomal population of a single cell.

Original languageEnglish (US)
Pages (from-to)4087-4092
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume115
Issue number16
DOIs
StatePublished - Apr 17 2018

Keywords

  • Electrochemical observation
  • Glucosidase activity
  • Nanocapillary
  • Single lysosomes
  • Subcellular analysis

ASJC Scopus subject areas

  • General

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