Fig. 2.

GSNOR colocalizes with NOS1 and RyR2 in the heart. (A) Confocal fluorescent microscopy images depict colocalization of GSNOR with NOS1 but not NOS3 and colocalization of GSNOR with RyR2 along the T-tubular invaginations of the cardiac myocyte. (Scale bars: 10 μM.) (B) GSNOR binds NOS1 but not NOS3 in vitro. Aliquots of recombinant NOS1 and NOS3 were incubated with GST or GST-GSNOR fusion proteins. The first well corresponds to the positive control, being either NOS1 or NOS3 (2.5 ng). The second well corresponds to the first flow through the column after binding. Consecutive washes of the column were then performed before elution. The bottom gel shows the profile (silver staining) of elution of GST-GSNOR (∼67 kDa) in the NOS1 binding experiment. (C) GSNOR enzymatic activity in total skeletal and heart muscle homogenates and in purified SR preparations. GSNOR activity is enriched in SR of cardiac but not skeletal muscle.