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Updated: Aug 6, 2026

Analysis of Protein Folding, Transport, and Degradation in Living Cells by Radioactive Pulse Chase
Published on: February 12, 2019
Detection of a hidden folding intermediate in the focal adhesion target domain: Implications for its function and
Zheng Zhou1, Hanqiao Feng, Yawen Bai
1Laboratory of Biochemistry, Center for Cancer Research, NCI, NIH, Bethesda, Maryland 20892, USA.
Abstract:
The focal adhesion target (FAT) domain of focal adhesion kinase has a four-helix bundle structure. Based on a hydrogen exchange-constrained computer simulation study and some indirect experimental results, it has been suggested that a partially unfolded state of the FAT domain with the N-terminal helix unfolded plays an important role in its biological function. Here, using a native-state hydrogen exchange method, we directly detected an intermediate with the N-terminal helix unfolded in a mutant (Y925E) of the FAT domain. In addition, kinetic folding studies on the FAT domain suggest that this intermediate exists on the native side of the rate-limiting transition state for folding. These results provide more direct evidence of the existence of the proposed intermediate and help to understand the folding mechanism of small single domain proteins.
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