Open AccessRevisiting Mitochondrial pH with an Improved Algorithm for Calibration of the Ratiometric 5(6)-carboxy-SNARF-1 Probe Reveals Anticooperative Reaction with H+ Ions and Warrants Further Studies of Organellar pH
Author(s) -
Tomasz Michał Żurawik,
Adam Pomorski,
Agnieszka Belczyk-Ciesielska,
Grażyna Goch,
K. M. Niedźwiedzka,
Róża Kucharczyk,
Artur Krężel,
Wojciech Bal
Publication year - 2016
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0161353
Subject(s) - mitochondrion , analyte , ion , biophysics , calibration , chemistry , fluorescence , analytical chemistry (journal) , biochemistry , biology , physics , chromatography , organic chemistry , quantum mechanics
Fluorescence measurements of pH and other analytes in the cell rely on accurate calibrations, but these have routinely used algorithms that inadequately describe the properties of indicators. Here, we have established a more accurate method for calibrating and analyzing data obtained using the ratiometric probe 5(6)-carboxy-SNARF-1. We tested the implications of novel approach to measurements of pH in yeast mitochondria, a compartment containing a small number of free H + ions. Our findings demonstrate that 5(6)-carboxy-SNARF-1 interacts with H + ions inside the mitochondria in an anticooperative manner (Hill coefficient n of 0.5) and the apparent pH inside the mitochondria is ~0.5 unit lower than had been generally assumed. This result, at odds with the current consensus on the mechanism of energy generation in the mitochondria, is in better agreement with theoretical considerations and warrants further studies of organellar pH.