Backbone dynamics of escherichia coli thioesterase/ protease i: Evidence of a flexible active-site environment for a serine protease

Yao Te Huang, Yen Chywan Liaw, Vitaliy Ya Gorbatyuk, Tai Huang Huang*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

47 Citations (Scopus)


Escherichia coli thioesterase/protease I (TEP-I) is a member of a novel subclass of the lipolytic enzymes with a distinctive GDSLS motif. In addition to possessing thioesterase and protease activities, TEP-I also exhibits arylesterase activity. We have determined the 15N nuclear magnetic spin relaxation rates, R1 and R2, and the steady state 1H15N heteronuclear Overhauser effect, measured at both 11.74 T and 14.09 T, of (u-15N) TEP-I. These data were analyzed using model-free formalism (with axially symmetric rotational diffusion anisotropy) to extract the backbone dynamics of TEP-I. The results reveal that the core structure of the central β-sheet and the long α-helices are rigid, while the binding pocket appears to be rather flexible. The rigid core serves as a scaffold to anchor the essential loops, which form the binding pocket. The most flexible residues display large amplitude fast (ps/ns time-scale) motion and lie on one stripe whose orientation is presumed to be the ligand-binding orientation. We also detected the presence of several residues displaying slow (μs/ms time-scale) conformational exchanging processes. These residues lie around the binding pocket and are oriented perpendicularly to the orientation of the flexible stripe. Two of the putative catalytic triads, Ser10 and His157, and their neighbors show motion on the μs/ms timescale, suggesting that their slow motion may have a role in catalysis, in addition to their possible roles in ligand binding. The presence of a flexible substrate-binding pocket may also facilitate binding to a wide range of substrates and confer the versatile functional property of this protein.

Original languageEnglish
Pages (from-to)1075-1090
Number of pages16
JournalJournal of Molecular Biology
Issue number4
Publication statusPublished - 2001 Apr 6
Externally publishedYes


  • Enzyme catalysis
  • NMR relaxation
  • Protein dynamics
  • Serine protease
  • Thioesterase

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Molecular Biology


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