摘要
报道了鸭肝脂肪酸合成酶在胍变性过程中构象变化和活性变化的关系，首次验证了邹承鲁提出的酶活性部位柔性构象的理论适用于多功能复合酶，同时该酶变性及复性均可测定出多个阶段，且证明有无活性稳态酶存在。在低浓度盐酸胍溶液中该酶的全反应活性和其中两个还原部位单独活性被同步可逆抑制，随着胍浓度增高，出现不可逆失活且程度和速度均迅速提高，在０．５４ｍｏｌ／Ｌ胍中该酶全反应活性在１．５分钟内已有一半不可逆丧失，终态只余约５％活性。失活过程有快相和慢相。全反应的失活程度及速度均大大高于两个还原反应。全反应的快相失活中，还原反应仍保持大部分活性，此时稀释可恢复活性。慢相失活使全反应和两个还原反应均失活，此时简单稀释不能恢复活性。在０．７２ｍｏｌ／Ｌ胍中该酶的内源荧光开始出现峰位红移和峰强下降，紫外差光谱增大，约１至２ｍｏｌ／Ｌ胍处有一个台阶，２．１６ｍｏｌ／Ｌ以上胍中其内源荧光和紫外差光谱再次迅速变化，２．８８ｍｏｌ／Ｌ以上胍形成第二台阶，不再变化。ＣＤ谱的变化是类似的规律。该天然二聚体酶中的的２０％硫基可和ＤＴＮＢ极快反应，应为表面硫基。胍处理后表面巯基增加，至２．８８ｍｏｌ／Ｌ胍中全部ＳＨ均已可迅速和ＤＴＮＢ作用。该酶的多阶?
The denaturation and inactiVation of fatty acid synthase (FAS) fiom duck liver bygUaindine hydrochioride (GndHCl) solution of different concentlation was studied. The overall activity and two nduction activities of FAS were reversibly inhibital by low conCentlationof GndHCl (below 0.45mol/L). Inactivation of the enzyme folioed an elhance of GndHClconcentlation. Half of the ovelall reaction activity was lost within l.5 Ininuthe in 0.54Inol/LGndHO, and 5% achvity remained at the end of inachvation. The inactivation involved twopad. The inactiVation of overan reaction activity was much mere breater and fastef than theinactiVation of reductions in the same condition and GndHCI conCentration. The overed reaction activity could be recovered by dilution ac fast pad inactiVatal by GndllCI, but not after second phase inactiVation in that phase the nductions were also inactivatal. The adults suggest that there are some stable unactive confonnations in the enzyme.The incease of UV absorbance of FAS, decrease in intheity and red shift of its intrinicfluorescence in GndHCl occur in two stages from 0.72 to 1.08mol/L and 2.16to 2.88mol/LGndHCl aspectively. A similarity was shown for its CD change. About 20% of the totalthiols of intaCt FAS natal rapidly with 5,5' -dithiobis (2-nitrobenzoic acid), and are believed to be suiface thiols . The total SH reacted rapidly with DTNB for the 2.88mol/LGndHCl treated FAS. Ihactivatoin occuring at a lower GndHCl concentration than that required for notiasble conformation changes suggests that the achve site of FAS is located in alimited region more fragile than the molecule as a whole.
出处
《生物物理学报》
CAS
CSCD
北大核心
1994年第1期6-12,共7页
Acta Biophysica Sinica
基金
国家教委优秀年轻教师基金
关键词
胍变性
构象变化
分子析叠
脂肪酸合成酶
失活化
Densturation by guandine Chnformation change Inactivation Molecular folding Fatty acid synthase
