S-ovalbumin, an ovalbumin conformer with properties analogous to those of loop-inserted serpins.
S-ovalbumin, an ovalbumin conformer with properties analogous to those of loop-inserted serpins.
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S-卵清蛋白，一种卵清蛋白构象异构体，具有与插入环的丝氨酸蛋白酶抑制剂类似的性质。
DOI：
10.1002/pro.5560040403
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Gettins,PG
中科院分区：
文献类型：
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作者：
Huntington,JA;Patston,PA;Gettins,PG
大多数丝氨酸蛋白酶抑制剂是丝氨酸蛋白酶的抑制剂，并且被认为在与蛋白酶形成复合物时经历构象变化，其涉及反应中心环部分插入抑制剂的β折叠中。卵清蛋白虽然是丝氨酸蛋白酶抑制剂，但不是丝氨酸蛋白酶的抑制剂。有人提出，这种缺陷是由于在反应中心区域的临界点（P14）存在带电残基精氨酸，这阻止了环插入β折叠，从而排除了抑制特性。为了测试在卵清蛋白中是否阻止环插入，我们检查了两种形式的卵清蛋白的性质：天然蛋白和S-卵清蛋白，S-卵清蛋白是从天然卵清蛋白自发形成的一种形式，具有增加的稳定性。量热测量表明S-卵清蛋白比卵清蛋白稳定约3 kcal mol−1。CD光谱表明S-卵清蛋白的α-螺旋比天然卵清蛋白少，1H NMR光谱表明总体结构非常相似，表明两种形式之间的构象差异有限。通过比较每种蛋白质的反应中心区域对猪胰弹性蛋白酶和枯草杆菌蛋白酶Carlsberg的蛋白水解的敏感性，我们得出结论，有限的天然S构象变化特别影响反应中心区域。这些数据与S-卵清蛋白的结构一致，其中部分反应中心环插入β-折叠A中以产生更稳定的结构，类似于其他丝氨酸蛋白酶抑制剂。然而，环插入的速率似乎比抑制性丝氨酸蛋白酶抑制剂低得多。因此，虽然环插入似乎没有被阻止的精氨酸在P14的存在下，它发生在太慢的速度有效地与底物途径竞争。仅这一点就足以说明卵清蛋白中缺乏可检测的抑制特性。
Most serpins are inhibitors of serine proteinases and are thought to undergo a conformational change upon complex formation with proteinase that involves partial insertion of the reactive center loop into a β‐sheet of the inhibitor. Ovalbumin, although a serpin, is not an inhibitor of serine proteinases. It has been proposed that this deficiency arises from the presence of a charged residue, arginine, at a critical point (P14) in the reactive center region, which prevents loop insertion into the β‐sheet and thereby precludes inhibitory properties. To test whether loop insertion is prevented in ovalbumin we have examined the properties of two forms of ovalbumin: the native protein and S‐ovalbumin, a form that forms spontaneously from native ovalbumin and has increased stability. Calorimetric measurements showed that S‐ovalbumin was more stable than ovalbumin by about 3 kcal mol−1. CD spectra, which indicated that S‐ovalbumin had less α‐helix than native ovalbumin, and1H NMR spectra, which indicated very similar overall structures, suggest limited conformational differences between the two forms. From comparison of the susceptibility of the reactive center region of each protein to proteolysis by porcine pancreatic elastase and by subtilisin Carlsberg, we concluded that the limited native‐to‐S conformational change specifically affected the reactive center region. These data are consistent with a structure for S‐ovalbumin in which part of the reactive center loop has inserted into β‐sheet A to give a more stable structure, analogously to other serpins. However, the rate of loop insertion appears to be very much lower than for inhibitory serpins. Thus, although loop insertion does not appear to be prevented by the presence of arginine at P14, it occurs at too slow a rate to compete effectively with the substrate pathway. This alone is sufficient to account for the absence of detectable inhibitory properties in ovalbumin.