Table 2.
Conversion of human and rat proinsulin in pulse-chased rat islet cells
Adenovirus | Proinsulin Conversion (%)
|
||
---|---|---|---|
Adeno-proins.wt | Adeno-proins.mut | ||
Proinsulin species | Human wt | Rat | Rat |
1 h Basal medium | 55.4 ± 7.8 | 35.1 ± 2.4 | 46.8 ± 8.5 |
+ 1 h stimulated medium | 96.5 ± 1.2 | 96.7 ± 0.1 | 97.9 ± 0.2 |
1 + 1 h cell extracts | 91.3 ± 3.2 | 94.7 ± 0.2 | 93.2 ± 0.1 |
6 h Basal medium | 70.7 ± 8.3 | 64.3 ± 4.5 | 72.8 ± 2.7 |
+ 1 h stimulated medium | 100 | 99.2 ± 0.1 | 99.5 ± 0.1 |
6 + 1 h cell extracts | 96.7 ± 0.6 | 97.8 ± 0.5 | 97.7 ± 0.4 |
mut, Arg/Gly32:Lys/Thr64 human proinsulin; wt, wild-type human proinsulin.
Islet cells were infected as indicated with adenovirus expressing wild-type (Adeno-proins.wt) or mutant (Arg/Gly32:Lys/Thr64) proinsulin (Adeno-proins.mut) and pulse labeled for 30 min. The cells were then chased under basal conditions for 1 or 6 h followed in each instance by a 1-h period of stimulation with secretagogues. Radioactive products were analyzed by HPLC. Proinsulin conversion is presented as (insulin/insulin + conversion intermediates + proinsulin) × 100. Data are for the conversion of human wild-type proinsulin or endogenous rat proinsulin in cells infected with one or the other recombinant adenovirus. There was no detectable conversion of the mutant proinsulin at any time (our unpublished data).