title: Inter-subunit cross-linking suppressed the dynamic oligomeric dissociation of Mycobacterium tuberculosis Hsp16.3 and reduced its chaperone activity.
abstract: Small heat shock proteins (sHsps) usually exist as dynamic oligomers and oligomeric dissociation was believed to be a prerequisite for their chaperone activities. The truth of this hypothesis was verified in our present study on Hsp16.3, one member of sHsps from Mycobacterium tuberculosis, mainly by utilizing chemical cross-linking. Analysis using size exclusion chromatography demonstrated that the heat-induced oligomeric dissociation of Hsp16.3 was severely blocked due to highly efficient inter-subunit cross-linkages generated by chemical cross-linking, as well as its chaperone activity being reduced. Further analysis by non-denaturing pore gradient polyacrylamide gel electrophoresis and fluorescence spectrometry revealed that the dynamic oligomeric dissociation/reassociation process of Hsp16.3 at room temperature was suppressed by inter-subunit cross-linkages, accompanied by significantly decreased exposure of hydrophobic surfaces that are usually hidden in oligomers. These findings supported the hypothesis that substrate-binding sites of sHsps are exposed presumably by dissociation of larger oligomers into smaller active oligomers, and therefore such a dissociation process could be adjusted to modulate chaperone activities.
authors.affiliation: Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, 100084, P. R. China.
authors.forename: Xinmiao
authors.initials: X
authors.lastname: Fu
authors.forename: Wangwang
authors.initials: W
authors.lastname: Jiao
authors.forename: Abuduaini
authors.initials: A
authors.lastname: Abulimiti
authors.forename: Zengyi
authors.initials: Z
authors.lastname: Chang
mesh_terms: D000817:Anilino Naphthalenesulfonates; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D018833:Chaperonins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002850:Chromatography, Gel; D002942:Circular Dichroism; D015180:Electrophoresis, Gel, Two-Dimensional; D009169:Mycobacterium tuberculosis / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
chemical_list: D000817:Anilino Naphthalenesulfonates; D001426:Bacterial Proteins; C431133:Hsp16.3 protein, Mycobacterium tuberculosis; D018833:Chaperonins