Skip to main content
Figure 2 | BMC Biochemistry

Figure 2

From: Lipid phosphate phosphatases dimerise, but this interaction is not required for in vivo activity

Figure 2

Immunoprecipitation assays (a) Cell lysates co-transfected with WunM3 and WunGFP (first 3 lanes of each gel) or WunD2M3 and WunGFP (lanes 4, 5 and 6). 'Load' is unbound protein removed from the resin after the 2 hour incubation. 'Wash' is an aliquot of the final buffer change. 'Capture' is an aliquot of the resin. This shows capture of WunM3 and WunD2M3 by the resin in the left hand blot. In the right hand blot, however, WunGFP is only detected in the lane corresponding to the WunM3 capture: there is no WunGFP detectable in the lane corresponding to the WunD2M3 capture. This indicates an association between full length Wun that is abolished by the truncation. The lane from cells transfected with only WunGFP (lane 7) shows that the resin does not capture the GFP tag. (b) 1% triton X-100 breaks the association – WunM3 no longer pulls down WunGFP. (c) mLPP-1M3 is captured and pulls down mLPP-1GFP indicating an association. This is not seen between WunM3 and hLPP-3GFP, or mLPP-1M3 and WunGFP (d). TL refers to 'total lysate' – an aliquot of the complete lysate prior to incubation with the resin. (e) WunD:248>TM3 does not pull down WunGFP. (f) Incubation of WunM3 or WunGFP in PFA indicates monomers (M) at 39 KDa/51 KDa and dimers (D) at 70 KDa/100 KDa plus higher order oligomers. TL = total lysate not treated with PFA. (g) WunD2GFP and WunD:248>TGFP do not form oligomers with PFA, whereas hLPP-3GFP and mLPP-1M3 (h) do.

Back to article page