Journal:

Article Title: Systematic Mutagenesis of the Murine Gammaherpesvirus 68 M2 Protein Identifies Domains Important for Chronic Infection ▿

doi: 10.1128/JVI.02234-07

Figure Lengend Snippet: Expression of mutant M2 proteins in mammalian cells. (A) Schematic illustration of the locations of PXXP motifs and tyrosine residues in the M2 protein which have been targeted for mutation in this report. (B) Cos-1 cells were transiently transfected with plasmids expressing the indicated forms of mutant M2 protein. At 48 hours posttransfection, cells were harvested and M2 immunoprecipitations were performed as described in Materials and Methods. (C) Cos-1 cells were transiently transfected with plasmids expressing the indicated forms of mutant M2 protein and GFP. At 24 hours posttransfection, 50 μg/ml CHX (+) or a vehicle control (−) was added to each sample, and cell lysates were harvested at the indicated times, in hours, posttreatment. Protein half-life was calculated by determining the time point at which the band intensity ratio of M2 to GFP was 50% of this ratio at time zero. WT, wt M2; Cntl, vector control; α, anti; IP, immunoprecipitation; t1/2, half-life.

Article Snippet: For M2 immunoprecipitations, affinity-purified chicken polyclonal anti-M2 antibodies (Gallus Immunotech, Ontario, Canada) were generated against the peptides GGRPQRPPLPRTRFC and CSTRSLMMLDSG, corresponding to amino acids 82 to 94 and 150 to 161, respectively.

Techniques: Expressing, Mutagenesis, Transfection, Control, Plasmid Preparation, Immunoprecipitation