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MMP-12是癌症治疗药物靶标.为了更好研制新药,需要大量制备MMP-12,但MMP-12在大肠杆菌中以包涵体形式表达.因此如何优化蛋白复性过程是大量获取MMP-12蛋白的关键.采用核磁共振、稳态荧光法、外源性ANS（8-anilinol-naphthalenesulfonic acid）荧光探针三种方法监控MMP-12变性蛋白的再折叠过程,以探究其复性折叠机制.研究发现MMP-12再折叠中点值与对应的尿素浓度几乎相等（Cm≈4,mid-point of transition）.不同尿素浓度中MMP-12的二维1H-15NHSQC（heteronuclear single quantum correlation）谱图显示,尿素浓度从4mol/L降低到3mol/L是MMP-12蛋白复性折叠的关键步骤.据此我们将MMP-12蛋白复性从常规的梯度透析复性方法改进成等容透析复性法（即确保尿素从4mol/L到3mol/L的浓度变化缓慢）,实现复性收率提高一倍.“,”MMP-12 is a drug target for cancer therapy,but it＇s over-expressed as inclusion bodies in E.coli To produce it in a large scale,in this paper,NMR,intrinsic tryptophan fluorescence,and ANS（8-anilinol-naphthalenesul fonic acid） fluorescence were employed to monitor its aggregation state to im-prove the refolding yield from inclusion bodies.The midpoint for the refolding（Cm≈4,mid-point of transi-tion） of MMP-12 obtained from NMR experiments coincides with those from intrinsic tryptophan fluores-cence and ANS fluorescence.These data suggest that the urea concentration from 4 mol/L to 3 mol/L is a key step in refolding process of MMP-12.Based on these results,we changed the refolding method from conventional step-wise dialysis to continuous dialysis,thus the refolding yield of MMP-12 from its inclusion bodies was doubled.