Fig. 6.

Characterization of Gln-299 mutants. (A) Chloride dependence of transport. Gluconate was used to replace Cl⁻ for concentrations below 200 mM with [Na⁺] constant at 200 mM. The graph shows a representative experiment. All error bars represent SDs from three measurements. The K_m value for Cl⁻ with WT was 13.56 ± 1.70 mM. This value represents the mean ± SEM of three experiments. (B) Gln299 mutants influence GlyT1b conformation and its response to Na⁺ and Cl⁻. Rate constants were determined for MTSET modification of Cys244 in the cytoplasmic pathway of S244C/C62A and Gln299 mutants in the S244C/C62A background. NMDG⁺ and gluconate were used as replacements for Na⁺ and Cl⁻, respectively. Ion concentrations were kept a constant at 150 mM in all experiments, and control represents the absence of NaCl. Control columns with * are significantly different from S244C/C62A, P < 0.01. Within each mutant, differences between control and Na⁺, or between Na⁺ and NaCl, where significant, are shown. *P < 0.01, using Student’s t test. All error bars represent the SEM; n ≥ 3.