K_{ATP }channels contain four Kir6.2 subunits. Macroscopic inside-out patch currents in response to voltage ramps (300 ms) between +100 mV (or *c*, +150 mV) and −100 mV, plotted versus membrane potential, for K_{ATP} channels formed from wild-type Kir6.2 (*a*, *WT*) or Kir6.2[N160D] (*b, N160D*) coexpressed with SUR1 subunits (in 1:1 molar ratio), or from mixed wild-type Kir6.2 and Kir6.2[N160D], plus SUR1 (*c*). (*c, top*) Currents are shown in the presence and absence of 20 μM spermine (*spm*), in each case after subtraction of leakage currents in the presence of 5 mM ATP (which reduces channel open probability to <1%). (*c, bottom*) Relative conductance (*G*_{REL})-voltage relationships from the data shown above. The smooth lines in *a* (and *b*) are best fits of single Boltzmann functions to the data. For these patches, the unconstrained Boltzmann functions (for i = 5 and 0, respectively) were fitted with *V*_{5} = +220 mV, *z*_{5} = 0.4 (*a*); *V*_{1} = −54 mV, *z*_{1} = 2.9 (*b*). In *c* (*bottom*), the smooth line that superimposes on the data is the sum of five Boltzmann functions that correspond to different combinations of wild-type Kir6.2 and Kir6.2[N160D] in a tetramer, with fitted probability of wild-type incorporation (*P*) = 0.56. For this patch, the unconstrained Boltzmann functions (for i = 2–4) components, were fitted with *V*_{2} = −48 mV, *z*_{2} = 2.9; *V*_{3} = +9 mV, *z*_{3} = 2.9; *V*_{4} = +86 mV, *z*_{4} = 1.0. The individual Boltzmann functions are also plotted, with the contributing number of wild-type and Kir6.2[N160D] subunits indicated beside each curve. (*d*) Residual errors (calculated as the sum of squares of the difference between the fit and the data, every 1 mV, between −80 to +160 mV, excluding −10 to +10 mV), obtained by fitting with *n* = 2–6 subunits [Σ(G_{REL} − G_{F})^{2}, where G_{F} is the fitted G_{REL}] plotted against the number of assumed subunits (*N*). The dashed line and open symbols correspond to the patch illustrated in *c*. Solid lines and symbols correspond to averaged data for *n* = 3–5 subunit fits for all patches (*n* = 9 patches, mean ± SEM). The trimeric model (*n* = 3) is inadequate to fit the data (see *d*), and better fits are not obtained by increasing *n* from 4 to 5. Similar results were obtained when attempting to fit the model to derivative (ΔG_{REL}/ΔV) data (not shown).

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