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Difference between revisions of "Respiratory acceptor control ratio"

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==Discussion==
== Discussion ==
ย 
The RCR is discussed in the context of [[OXPHOS]] versus [[ETS capacity]].
The RCR is discussed in the context of [[OXPHOS]] versus [[ETS capacity]].
== Problem: The RCR is low in isolated mitochondria ==
At a high mitochondrial concentration (0.5 mg protein/ml), ADP is exhausted very rapidly if added at a concentration typically used for [[State 3]] measurements (200-300 ยตM), which may stimulate flux below the ADP-saturated state of [[OXPHOS capacity]]. The slope increases sharply and returns to [[State 4]] ([[LEAK]]), even before the plot of flux shows the true maximum, resulting in a low apparent '''[[RCR]]'''.
=== Tests and Solutions ===
# Reduce the data recording interval (standard is 2 seconds) to the minimum of 0.2 s (setting in the Oxygraph Control window; see [[MiPNet12.06 O2k-Start]]). As the data recording interval is reduced, the flux appears more noisy, but represents transitions more accurately and reduces the apparent time-delay. At high flux per volume, the increased noise level presents no problem. Flux then shows a period of constant OXPHOS capacity rather than a sharp peak.
# Reduce the mitochondrial concentration, thus prolonging the duration of ADP-saturated respiration (OXPHOS capacity) over 120 seconds. If flux reaches a constant maximum value, then analysis is possible without application of signal deconvolution (standard plot of flux in DatLab).
# Otherwise, export the data into [http://www.oroboros.at/?O2k-datlab2 DatLab 2], and apply a time correction (signal deconvolution), as described by [[Gnaiger 2001 Respir Physiol]].
# See also โ€˜Notes on [[Time resolution]]โ€™.

Revision as of 12:46, 5 October 2012


high-resolution terminology - matching measurements at high-resolution


Respiratory acceptor control ratio

Description

The respiratory control ratio (RCR) is defined as State 3/State 4 (Chance and Williams 1955). Considering an index of uncoupling, RCR should be replaced by the L/E ratio, if P/E<1.0 (Gnaiger 2009). In intact cells, RCR has been used for the ratio State 3u/State 4o, i.e. for the inverse L/E ratio (Huetter et al., 2004).

Abbreviation: RCR

Reference: Chance 1955 JBC-III; MiPNet12.15


MitoPedia methods: Respirometry 


MitoPedia topics: "Respiratory state" is not in the list (Enzyme, Medium, Inhibitor, Substrate and metabolite, Uncoupler, Sample preparation, Permeabilization agent, EAGLE, MitoGlobal Organizations, MitoGlobal Centres, ...) of allowed values for the "MitoPedia topic" property. Respiratory state"Respiratory state" is not in the list (Enzyme, Medium, Inhibitor, Substrate and metabolite, Uncoupler, Sample preparation, Permeabilization agent, EAGLE, MitoGlobal Organizations, MitoGlobal Centres, ...) of allowed values for the "MitoPedia topic" property. 

Discussion

The RCR is discussed in the context of OXPHOS versus ETS capacity.


Problem: The RCR is low in isolated mitochondria

At a high mitochondrial concentration (0.5 mg protein/ml), ADP is exhausted very rapidly if added at a concentration typically used for State 3 measurements (200-300 ยตM), which may stimulate flux below the ADP-saturated state of OXPHOS capacity. The slope increases sharply and returns to State 4 (LEAK), even before the plot of flux shows the true maximum, resulting in a low apparent RCR.

Tests and Solutions

  1. Reduce the data recording interval (standard is 2 seconds) to the minimum of 0.2 s (setting in the Oxygraph Control window; see MiPNet12.06 O2k-Start). As the data recording interval is reduced, the flux appears more noisy, but represents transitions more accurately and reduces the apparent time-delay. At high flux per volume, the increased noise level presents no problem. Flux then shows a period of constant OXPHOS capacity rather than a sharp peak.
  2. Reduce the mitochondrial concentration, thus prolonging the duration of ADP-saturated respiration (OXPHOS capacity) over 120 seconds. If flux reaches a constant maximum value, then analysis is possible without application of signal deconvolution (standard plot of flux in DatLab).
  3. Otherwise, export the data into DatLab 2, and apply a time correction (signal deconvolution), as described by Gnaiger 2001 Respir Physiol.
  4. See also โ€˜Notes on Time resolutionโ€™.