Identification of a potential hydrophobic peptide binding site in the C‐terminal arm of trigger factor
Identification of a potential hydrophobic peptide binding site in the C‐terminal arm of trigger factor
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触发因子 C 末端臂中潜在疏水性肽结合位点的鉴定
DOI：
10.1110/ps.062623707
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发表时间：
2007
期刊：
影响因子：
8
通讯作者：
Jun‐mei Zhou
中科院分区：
文献类型：
--
作者：
Yi Shi;Dong;Shuxiang Li;Hong‐jie Zhang;S. Perrett;Jun‐mei Zhou
Trigger factor (TF) is the first chaperone to interact with nascent chains and facilitate their folding in bacteria. Escherichia coli TF is 432 residues in length and contains three domains with distinct structural and functional properties. The N‐terminal domain of TF is important for ribosome binding, and the M‐domain carries the PPIase activity. However, the function of the C‐terminal domain remains unclear, and the residues or regions directly involved in substrate binding have not yet been identified. Here, a hydrophobic probe, bis‐ANS, was used to characterize potential substrate‐binding regions. Results showed that bis‐ANS binds TF with a 1:1 stoichiometry and a Kd of 16 μM, and it can be covalently incorporated into TF by UV‐light irradiation. A single bis‐ANS–labeled peptide was obtained by tryptic digestion and identified by MALDI‐TOF mass spectrometry as Asn391‐Lys392. In silico docking analysis identified a single potential binding site for bis‐ANS on the TF molecule, which is adjacent to this dipeptide and lies in the pocket formed by the C‐terminal arms. The bis‐ANS‐labeled TF completely lost the ability to assist GAPDH or lysozyme refolding and showed increased protection toward cleavage by α‐chymotrypsin, suggesting blocking of hydrophobic residues. The C‐terminal truncation mutant TF389 also showed no chaperone activity and could not bind bis‐ANS. These results suggest that bis‐ANS binding may mimic binding of a substrate peptide and that the C‐terminal region of TF plays an important role in hydrophobic binding and chaperone function.
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影响因子：
2.9
作者：
Prasad,AR;Luduena,RF;Horowitz,PM
通讯作者：
Horowitz,PM
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影响因子：
--
作者：
Seale,JW;Brazil,BT;Horowitz,PM
通讯作者：
Horowitz,PM
影响因子：
2.9
作者：
A. L. Smoot;M. Panda;Bill T. Brazil;A. Buckle;and Alan R. Fersht;P. Horowitz
通讯作者：
A. L. Smoot;M. Panda;Bill T. Brazil;A. Buckle;and Alan R. Fersht;P. Horowitz
影响因子：
2.9
作者：
PRASAD, ARS;LUDUENA, RF;HOROWITZ, PM
通讯作者：
HOROWITZ, PM
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影响因子：
2.9
作者：
Seale,JW;Martinez,JL;Horowitz,PM
通讯作者：
Horowitz,PM