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Updated: May 14, 2026

Combining Chemical Cross-linking and Mass Spectrometry of Intact Protein Complexes to Study the Architecture of Multi-subunit Protein Assemblies
Published on: November 28, 2017
Interactions between subunits a and b in the rotary ATP synthase as determined by cross-linking
Jessica DeLeon-Rangel1, Robert R Ishmukhametov, Warren Jiang
1Department of Biological Sciences, Southern Methodist University, Dallas, TX 75275-0376, USA.
Abstract:
The interaction of the membrane traversing stator subunits a and b of the rotary ATP synthase was probed by substitution of a single Cys into each subunit with subsequent Cu(2+) catalyzed cross-linking. Extensive interaction between the transmembrane (TM) region of one b subunit and TM2 of subunit a was indicated by cross-linking with 6 Cys pairs introduced into these regions. Additional disulfide cross-linking was observed between the N-terminus of subunit b and the periplasmic loop connecting TM4 and TM5 of subunit a. Finally, benzophenone-4-maleimide derivatized Cys in the 2-3 periplasmic loop of subunit a were shown to cross-link with the periplasmic N-terminal region of subunit b. These experiments help to define the juxtaposition of subunits b and a in the ATP synthase.
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