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. 2019 Jan 3;8:e39180. doi: 10.7554/eLife.39180

Figure 3. The intrinsic PDZ affinity does not translate directly to avidity but determines maximal binding.

(b) Normalized binding derived from SCMS as a function of PICK1 concentration to LKV (black) (Kd*=37 ± 5 nM, Bmax = 100%), LKI (dark grey) (Kd*=29 ± 4 nM, Bmax = 56 ± 2%), LKA (light grey) (Kd*=59 ± 11 nM, Bmax = 41 ± 4%) LKV +A (white) (Kd*not fitted, Bmax = 21 ± 3). n = 8, 7, 5 and 3, respectively (**p<0.01; ****p≤0.0001). (b) Representative images demonstrating concentration dependent binding of PICK1 to SCMS expressing LKV, LKI and LKA constructs. (c) Representative PICK1 binding to SCMS expressing LKV or LKI as a function of incubation time. PICK1 concentration is 100 nM. Half maximum binding values are 24 ± 14 min for LKV, and 11 ± 4 min for LKI (means ±s.e.m, n = 3). (d) Representative images showing time dependent PICK1 binding to LKV and LKI. (e) Representative PICK1 dissociation curves from SCMS expressing LKV or LKI (points are means ±SD). LKV is fitted to a two-state dissociation with estimated fast and slow half-life of 21 ± 8 and 373 ± 51 min., respectively. LKI is fitted to a one-state dissociation with a half-life of 431 ± 16 min. (means ± S.E, n = 3). (f) Representative images showing time dependent PICK1 dissociation from LKV and LKI. Scale bars 10 μm.

Figure 3.

Figure 3—figure supplement 1. β2 fusion constructs and expression levels.

Figure 3—figure supplement 1.

(a) Schematic representation of β2 C-terminal fusion constructs. (b) Expression levels of β2 DAT fusion constructs. Secondary units are arbitrary units relating to measured intensities.