Assembly Mechanism And Folding Chaperones Identification Of C-reactive Protein

Natural human C-reactive protein(CRP),composed of five identical subunits which associate each other into a disc-like structure,is a prototypic acute phase protein which is mainly secreted by the liver.CRP concentration in serum increased more than 1000-fold from 1?g/ml baseline level within 48 hours after onset of tissue injury and inflammation.CRP is used to be a marker to evaluate and monitor the development of infection,tissue injury and inflammation related diseases,especially in cardiovascular disease.Emerging evidence in vitro indicate that CRP not only a marker,but also a direct mediator through conformation dissociation to form monomeric CRP.However,contradiction between pCRP and mCRP remained regarding their activities in vivo because of complexity added by CRP conformation changing.To research specific role of CRP in pathophysiology,it is necessary and meaningful to research CRP folding,assembly and secretory for constructing animal models which only express one type of CRP conformation.Wild type CRP only secreted in pentamer form from cells,which is similar to CRP secretion in vivo.The phenomenon indicate assembly of CRP subunit is necessary to its secretion and monitored by cell quality control system.Mutant Y40C/V117 C formed pantamer and be secreted from cells.Further analysis revealed that the mutant formed inter-subunit disulfide bond and mean while the native disulfide bond in subunit was remained.This result indicate subunit folding precedes pentamer assembly and assembly of CRP subunit start only when its structure is similar to native subunit conformation.The conclusion fit the evidence of pCRP dissociation in urea denaturing solution which reflected pentameric CRP depend on the third conformation of subunit.From the result of expression and protein interaction of mutant of disulfide bond,calcium site and ionic bond,we inferred the folding of CRP subunit include two main process.First,the peptide start folding from N-terminal and form hydrophobic core.When Cys36 and Cys97 in contact,the disulfide bond formed and stabilized hydrophobic core.After that,calcium took part in the following folding process and made the subunit more compact and similar to the native conformation.In process of subunit folding,DnaK and GroEL are main folding chaperones which could assisted the third structure formation of subunit.But at this time,the subunit assembly cannot occur and still need the help of peptidyl-prolyl cis-trans isomerase which made the subunit more contact by the isomerization of conservative proline and fine-tuning of local region.Then,assembly of pentamer may occur.