title: Small heat shock protein IbpB acts as a robust chaperone in living cells by hierarchically activating its multi-type substrate-binding residues.
abstract: As ubiquitous molecular chaperones, small heat shock proteins (sHSPs) are crucial for protein homeostasis. It is not clear why sHSPs are able to bind a wide spectrum of non-native substrate proteins and how such binding is enhanced by heat shock. Here, by utilizing a genetically incorporated photo-cross-linker (p-benzoyl-l-phenylalanine), we systematically characterized the substrate-binding residues in IbpB (a sHSP from Escherichia coli) in living cells over a wide spectrum of temperatures (from 20 to 50 °C). A total of 20 and 48 residues were identified at normal and heat shock temperatures, respectively. They are not necessarily hydrophobic and can be classified into three types: types I and II were activated at low and normal temperatures, respectively, and type III mediated oligomerization at low temperature but switched to substrate binding at heat shock temperature. In addition, substrate binding of IbpB in living cells began at temperatures as low as 25 °C and was further enhanced upon temperature elevation. Together, these in vivo data provide novel structural insights into the wide substrate spectrum of sHSPs and suggest that sHSP is able to hierarchically activate its multi-type substrate-binding residues and thus act as a robust chaperone in cells under fluctuating growth conditions.
authors.affiliation: From the State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Center for Protein Sciences, Peking University, Beijing 100871, China.
authors.forename: Xinmiao
authors.initials: X
authors.lastname: Fu
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Shi
authors.forename: Linxiang
authors.initials: L
authors.lastname: Yin
authors.forename: Jiafeng
authors.initials: J
authors.lastname: Liu
authors.forename: Keehyoung
authors.initials: K
authors.lastname: Joo
authors.forename: Jooyoung
authors.initials: J
authors.lastname: Lee
authors.forename: Zengyi
authors.initials: Z
authors.lastname: Chang
mesh_terms: D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D018869:Heat-Shock Response / Q000502:physiology*; D006358:Hot Temperature; D010449:Peptide Mapping / Q000379:methods; D055503:Protein Multimerization / Q000502:physiology*
chemical_list: D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; C097579:IbpB protein, E coli