FtsH3 + FtsH10 | ATP-dependent zinc metalloprotease FtsH3 + FtsH10 (mitochondrial)
AS07 204 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Arabidopsis thaliana

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Product Information
KLH-conjugated peptide dereived from sequences of Arabidopsis thaliana FtsH3 and FtsH10 with localization to mitochondria Q84WU8, At2g29080 and Q8VZI8, At1g07510
80 kDa
Reactivity
Arabidopsis thaliana
Application examples
Application example

Total protein from Arabidopsis thaliana mitochondria (20 µg) were separated on 10% acrilamide gel and electrophoresis prepared according to Schägger and von Jagov (Anl. Biochem., 1987, 166:368-379). After running the gel, proteins were transferred to nitrocellulose membrane using wet transfer (0.22% CAPS, pH 11). Transfer was checked by Ponceau S staining. Blot was destained by several quick washings in distilled water and 1 washing in 1X TBS (10 mM T pH 7.5, 150 mM NaCl) (10-15 min.).Blot was blocked by 1.5 hour in 5% milk in TBST (1X TBS, 0,1 20) After blocking blot was washed quickly twice in TBST and incubated 2 hours with primary antibody (dilution 1: 1000 TBST (dilution 1:1000). Washing: two quick washings in TBST and 3 x 10 min. washings in TBST. Then blot was incubated 45-60 min. with a secondary anti-rabbit antibodies conjugated to peroxidase (dilution 1:10000) in TBST. Washing: as above. After washing blot was incubated 1-2 min. in ECL solution and exposed to Kodak autoradiography film. Exposure time was 15-60 seconds.
Mitochondria were isolated as described by Urantowka et al. (Plant Mol Biol, 2005, 59:239-52). Mitochondrial pellets were suspended in 1X Laemmli buffer (5% beta-mercaptoetanol, 3.7% glycerol, 1.1% SDS, 23 mM Tris-HCl pH 6.8, 0.01% bromophenol blue), heated (95ºC, 5 min.) and centrifuged (13 000rpm, 1 min.).
Courtesy Dr. J. Piechota, Wrocław University, Poland
Additional information
Blue-native (2D BN/SDS-PAGE) methodology is described in Piechota et al. 2010
Background
One of the several classes of mitochondrial proteases is membrane bound, ATPdependent FtsH protease. Their function is very important for the control of protein quality and quantity by degradation of unassembled subunits. Other names: AtFtsH3, cell division protease ftsH homolog 3, mitochondrial, AtFtsH10, cell division protease ftsH homolog 10, mitochondrial
Product citations
Piechota et al. (2010). Identification and characetization of high-molecular-weight complexes fromed by m-AAA proteases and prohibitins in mitochondria of Arabidopsis thaliana. J Biol Chem. 2010 Apr 23;285(17):12512-21. doi: 10.1074/jbc.M109.063644.
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