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. 2009 Nov 23;285(5):3168–3180. doi: 10.1074/jbc.M109.056846

FIGURE 7.

FIGURE 7.

Protein S-glutathionylation of the 51-kDa subunit of NQR in myocardial tissue homogenates and the effect of post-ischemic injury. A, mitochondrial preparation (1 mg) from rat heart was subjected to immunoprecipitation with Ab51 antibody and subsequently subjected to SDS-PAGE (left-side panel) and immunoblotting with anti-GSH monoclonal antibody (right-side panel). M denotes a molecular weight marker standard. B, myocardial tissue homogenates (300 μg) were obtained from the infarction region (left-side panel, in vivo regional I/R rat heart) or isolated heart subjected to global I/R injury (right-side panel). Tissue homogenates were immunoprecipitated with Ab51 and then probed with anti-GSH antibody and Ab51. M denotes the molecular weight marker obtained from S-glutathionylated NDH at the 51-kDa FMN-binding subunit (12), C denotes non-ischemic tissue (left-side panel) or normal control rat heart (right-side panel). C, Ab75 was immobilized to Affi-Gel 10 in accordance with the product literature. Mitochondrial preparation (1 mg) from rat heart was subjected to immunoprecipitation with immobilized Ab75. Proteins obtained from immunoprecipitation were subjected to SDS-PAGE (left-side panel) and probed with anti-GSH monoclonal antibody (right-side panel). D, same as B, except that Ab75 was used for immunoprecipitation. The density ratio of the blotting signals between anti-GSH and Ab51 (in B) or Ab75 (in D) was quantitated by using ImageJ.