Calponin cDNA was site-mutated (Thr184→Ala184) by overlap extension PCR method to reduce the opportunity for calponin to be phosphorylated by PKC or CaMKⅡ. The PCR product was sequenced. The mutated cDNA was then recombined into an expression vector pAED₄ and transferred into, expressed in E.coli BD₂₁(DE₃) induced by IPTG. The expression level was up to 15% of total bacterial proteins. Mutated calponin was liberated form E.coli by freezing and thawing and purified by Sephadex G-50. The recombinant protein and the purified mutant protein was identified by Western blotting.