Expression Of Human C-Reactive Protein In Pichia Pastoris

C‐reactive protein (CRP) is an acute phase plasma protein produced by the liver. The biosynthesis of CRP and its circulating concentration dramatically increase in inflammatory states; however, the in vivo functions of CRP have not been defined. To investigate the functions of CRP employing animal models of inflammatory diseases, there is a need for an in vitro expression system from which CRP can be obtained in large quantity. We developed a eukaryotic expression system for CRP using the yeast Pichia pastoris . The cDNA encoding human CRP was codon‐optimized via gene synthesis for expression in P. pastoris . The protein was directed for secretion by the incorporation of yeast α‐mating factor as an amino‐terminal fusion domain that is removed during secretion. CRP was successfully expressed and could be isolated from the culture media by affinity chromatography based on the calcium‐dependent binding specificity of CRP for phosphocholine. The pentameric structure and the phosphocholinebinding activity of recombinant CRP were not different from that of native CRP. Further characterization, such as the complement‐activating function, of yeast‐expressed recombinant CRP is in progress. The yeast expression system produces endotoxin‐free CRP and is also beneficial for expressing various CRP mutants which can be used to investigate structure‐function relationships of CRP. NIH grants R15HL091770 and R01HL071233