Santos A M C, Santana M A, Gomide F T F, Miranda A A C, Oliveira J S, Boas F A S Vilas, Vasconcelos A B, Bemquerer M P, Santoro M M
Department of Biochemistry and Immunology, Biological Sciences Institute, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, MG, Brazil.
Int J Biol Macromol. 2008 Apr 1;42(3):278-84. doi: 10.1016/j.ijbiomac.2007.12.002. Epub 2007 Dec 23.
alpha-Trypsin is a serine-protease with a polypeptide chain of 223 amino acid residues and six disulfide bridges. It is a globular protein with predominance of antiparallel ss-sheet secondary structure and it has two domains with similar structures. In the present work, a stability study of alpha-trypsin in the acid pH range was performed and some physical-chemical denaturation parameters were measured by using differential scanning calorimetry (DSC). The alpha-trypsin has a shelf-life (t(95%)) of about 10 months at pH 3.0 and 4 degrees C and its hydrolysis into the psi-trypsin isoform is negligible during 6 months. The observed ratio DeltaH(cal)/DeltaH(vH) is close to unity, which suggests the occurrence of a two-state transition. At pH 3.0, alpha-trypsin unfolded with T(m) = 325.9 K and DeltaH = 99.10 kcal mol(-1), and the change in heat capacity between the native and unfolded forms of the protein was estimated to be 1.96+/-0.18 kcal mol(-1)K(-1). The stability of alpha-trypsin calculated at 298 K was DeltaG(U)=6.10 kcal mol(-1) at pH 3.0. These values are in the range expected for a small globular protein. These results show that the thermodynamic parameters of unfolding of beta-trypsin do not change substantially after its conversion to alpha-trypsin.
α-胰蛋白酶是一种丝氨酸蛋白酶，具有一条由223个氨基酸残基组成的多肽链和六个二硫键。它是一种球状蛋白质，以反平行β-折叠二级结构为主，有两个结构相似的结构域。在本研究中，对α-胰蛋白酶在酸性pH范围内的稳定性进行了研究，并使用差示扫描量热法（DSC）测量了一些物理化学变性参数。α-胰蛋白酶在pH 3.0和4℃下的保质期（t(95%)）约为10个月，在6个月内其水解为ψ-胰蛋白酶同工型的情况可忽略不计。观察到的ΔH(cal)/ΔH(vH)比值接近1，这表明发生了两态转变。在pH 3.0时，α-胰蛋白酶在T(m)=325.9 K和ΔH = 99.10 kcal mol(-1)时展开，蛋白质天然态和展开态之间的热容变化估计为1.96±0.18 kcal mol(-1)K(-1)。在298 K下计算的α-胰蛋白酶在pH 3.0时的稳定性为ΔG(U)=6.10 kcal mol(-1)。这些值在小球状蛋白质预期的范围内。这些结果表明，β-胰蛋白酶转化为α-胰蛋白酶后，其展开的热力学参数没有实质性变化。
