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Data S1. Supplemental methods.

Figure S1. Characterization of Munc13-1 C1.

Figure S2. Circular dichroism spectra of Munc13-1 C1 and its mutants: Comparison of circular dichroism spectra of Munc13-1 C1 (____), E582A (………), E582L (__ __ __), and E582H (_ _ _).

Figure S3. 3-azibutanol and 3-azioctanol quench the fluorescence of Munc13-1C1.

Figure S4. Glu-582 as the site for 3-azibutanol.

Figure S5. Identification of Glu-599 as the site for 3-azibutanol in E582A.

Figure S6. Effect of alcohols on thermal unfolding of wild-type and mutant protein (E582A).

Table S1. Photolabeling of Munc13-1 C1 mutants analyzed by mass spectrometry.

Table S2. Summary of melting temperatures (Tm) of Munc13-1 C1 and its mutants determined by circular dichroism.

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