FIGURE 3.
Mutant Kirrel2Y595F/Y596F protein displays enhanced stability. A, MIN6 cells expressing wild type or mutant Kirrel2-V5 were treated with pervanadate. V5-tagged proteins were immunoprecipitated (IP) with anti-V5-agarose affinity gels and immunoblotted (IB) with 4G10 and anti-V5 antibodies. B, MIN6 cells were transfected with IGF1R-V5, control (pcDNA), wild type, or mutant Kirrel2 vectors. Duplicate transfections were performed using independent maximum preparations. Total protein lysates were immunoblotted with indicated antibodies. γ-Tub, γ-tubulin. C, MIN6 cells, co-transfected with GFP and wild type or mutant Kirrel2-V5 expression vectors, were treated with cycloheximide for the indicated time. Cell extracts were immunoblotted with V5 tag and GFP antibodies. D, quantification of Kirrel-V5 signals from two independent experiments by densitometry. Two-way analysis of variance with Bonferroni post hoc test was performed. **, p < 0.01; ***, p < 0.001.