Interaction between SNAP-23 and syntaxin 6.

(A) In vitro binding of syntaxin 6 to GST-SNAP-23. Unfused GST and full-length GST–SNAP-23 fusion protein were immobilized on glutathione-Sepharose beads and incubated with Triton X-100–extracts of human neutrophils. Then, bound proteins were eluted, analyzed by SDS-polyacrylamide gel electrophoresis, and blotted with 3D10 anti–syntaxin 6 MoAb. Triton X-100 extract of human neutrophils was used as a positive control. The position of syntaxin 6 is indicated. The experiment shown is representative of 3 performed. (B) Coimmunoprecipitation of SNAP-23 and syntaxin 6 from resting and activated human neutrophils. SNAP-23 was immunoprecipitated from detergent-solubilized whole cell extracts of resting and PMA-activated human neutrophils. Immunoprecipitates were electrophoresed in SDS-polyacrylamide gels and immunoblotted with the anti–SNAP-23 or anti–syntaxin 6 antibodies. Lysates were also immunoprecipitated with P3X63 myeloma culture supernatant as a negative control (Control). The positions of SNAP-23 and syntaxin 6 are indicated. Experiments shown are representative of 3 performed.