P. Wiesner et al., ISOLATION OF MITOCHONDRIAL PORIN OF THE FLY PROTOPHORMIA - PORIN MODIFICATION BY THE PESTICIDE CGA-140'408 STUDIED IN LIPID BILAYER-MEMBRANES, Biochimica et biophysica acta. Biomembranes, 1282(2), 1996, pp. 216-224
Mitochondrial porin from the fly Protophormia was solubilized with det
ergent from whole mitochondria and purified by chromatography across a
hydroxyapatite (HPT) column. The purified protein had an apparent mol
ecular mass of about 30 kDa on SDS-PAGE. partial sequencing of the pro
tein confirmed that it is porin. When reconstituted in planar lipid bi
layer membranes, porin formed ion-permeable channels with single-chann
el conductances of 2.4 and 4.5 nS in 1 M KCI. At low voltage, Protopho
rmia porin displayed the properties of a general diffusion pore and ha
d a small selectivity for anions over cations, At transmembrane potent
ials starting with about 20-30 mV, the channel switched in closed stat
e, which is still ion-permeable. Our results suggest that Protophormia
porin possesses functional properties similar to these of other mitoc
hondrial porins. Porin was also isolated and purified from mitochondri
a, which were treated with the carbodiimide CGA 140'408 It represents
the active derivative of diafenthiuron a new acaricide and insecticide
. This carbodiimide labels both a F-0-component of the inner membrane
ATPase and outer membrane porin in a similar way as N,N'-dicyclohexylc
arbodiimide (DCCD). Reconstitution experiments with the CGA 140'408-mo
dified porin showed no significant effect of the modification on the s
ingle-channel conductance, suggesting that CGA 140'408 binds outside t
he channel. The voltage-dependence of the CGA 140'408-modified porin w
as changed with respect to the unmodified form, The closed configurati
on of the pesticide-modified channel was reached at smaller transmembr
ane potentials, suggesting a shift of the open to the closed state of
Protophormia porin by pesticide binding. A possible contribution of th
is effect to the pesticide action is discussed.