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. 2004 Aug;78(15):8172–8182. doi: 10.1128/JVI.78.15.8172-8182.2004

FIG. 6.

FIG. 6.

Kinetic analysis of PABP cleavage in FCV-infected cells. CRFK cells were infected with FCV as described previously, and cells were isolated at hourly time points (indicated above lanes) postinfection for analysis. (A) Immunoblot analysis of feline PABP over time in FCV-infected cells detected with antibodies raised against human PABP. An analysis of mock-infected cells harvested after 8 h incubation is shown (M). (B) Analysis of FCV-infected cell lysates in the in vitro assay for human PABP cleavage. Aliquots of infected cell extracts from time points 1 to 8 hpi were mixed with radiolabeled human PABP and incubated for 3.5 h at 30°C. Controls include radiolabeled PABP incubated with recombinant FCV ProPol enzyme, mock-infected cell extract (M), and buffer alone (C). Autoradiogram of SDS-polyacrylamide gel is shown. Arrowheads indicate PABP cleavage products. Migration of molecular weight markers is indicated on the right. The human PABP-specific antibody used in the immunoblot (A) is peptide-specific (sequence located in RRM4) and recognizes only the amino-terminal cleavage product of PABP.