Intra-subunit Disulfide Determines the Conversion and Structural Stability of CRP Isoforms

• Published in: Inflammation 2020-04, Vol.43 (2), p.466-477
• Main Authors: Zhang, Chun-Miao, Tan, Yu-Bo, Zhou, Hai-Hong, Ge, Zhong-Bo, Feng, Jun-Rui, Lv, Guang-Bo, Sun, Zhi-Yuan, Fu, Yu, Wang, Ming-Yu
• Format: Article
• Language: English
• Subjects: Biomedical and Life Sciences; Biomedicine; C-reactive protein; C-Reactive Protein - chemistry; C-Reactive Protein - ultrastructure; Circular dichroism; Disulfides - chemistry; Dose-Response Relationship, Drug; Electron microscopy; Electrostatic properties; Fluorescence spectroscopy; Humans; Immunology; Inflammation; Intermediates; Internal Medicine; Isoforms; Microscopy, Electron - methods; Microscopy, Fluorescence - methods; Original Article; Pathology; Pharmacology/Toxicology; Protein Isoforms - chemistry; Protein Isoforms - ultrastructure; Protein Stability - drug effects; Protein structure; Protein Subunits - chemistry; Rheumatology; Secondary structure; Spectrum analysis; Urea - pharmacology
• ISSN: 0360-3997; 1573-2576
• DOI: 10.1007/s10753-019-01130-x

C-reactive protein (CRP) is a major human acute-phase reactant that is composed of five identical subunits. CRP dissociates into subunits at inflammatory loci forming monomeric CRP (mCRP) with substantially enhanced activities, which can be further activated by reducing the intra-subunit disulfide bond. However, conformational changes underlying the activation process of CRP are less well understood. Conformational changes accompanying the conversion of CRP to mCRP with or without reduction were examined with circular dichroism spectroscopy, fluorescence spectroscopy, electron microscopy, size-exclusion chromatography, and neoepitope expression. The conversion of CRP to mCRP follows a two-stage process. In the first stage, CRP dissociates into molten globular subunits characterized by intact secondary structure elements with greatly impaired tertiary packing. In the second stage, these intermediates completely lose their native subunit conformation and assemble into high-order aggregates. The inclusion of reductant accelerates the formation of molten globular subunits in the first step and promotes the formation of more compact aggregates in the second stage. We further show a significant contribution of electrostatic interactions to the stabilization of native CRP. The conformational features of dissociated subunits and the aggregation of mCRP may have a key impact on their activities.