Protein/Ice Interaction: High-Resolution Synchrotron X-ray Diffraction Differentiates Pharmaceutical Proteins from Lysozyme.
Protein/Ice Interaction: High-Resolution Synchrotron X-ray Diffraction Differentiates Pharmaceutical Proteins from Lysozyme.
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DOI：
10.1021/acs.jpcb.9b02443
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发表时间：
2019-07-11
影响因子：
3.3
通讯作者：
Shalaev, Evgenyi
中科院分区：
文献类型：
--
作者：
Bhatnagar, Bakul;Zakharov, Boris;Fisyuk, Alexander;Wen, Xin;Karim, Fawziya;Lee, Kimberly;Seryotkin, Yurii;Mogodi, Mashikoane;Fitch, Andy;Boldyreva, Elena;Kostyuchenko, Anastasia;Shalaev, Evgenyi
虽然蛋白质通常可以通过将它们保持在冰冻状态来稳定，但水到冰的转变也会导致蛋白质分子的降解。本文提出了一种研究蛋白质/冰相互作用的新方法，该方法基于测量六角形冰(Ih)的X射线衍射图特征。在欧洲同步辐射装置的ID22光束线上，用高分辨率同步辐射X射线衍射仪研究了四种不同蛋白质和几种小分子溶质的水溶液。对光束线进行了优化，消除了衍射线的仪器展宽，减少了择优取向效应，从而能够对X射线衍射数据进行定量分析。分析表明，重组人白蛋白(RHA)和单抗(MAb)这两种药物蛋白对冰晶的性质有显著的影响。特别是，与模型蛋白(溶菌酶)、抗冻蛋白、蔗糖和组氨酸相比，药物蛋白溶液中结晶结构域的尺寸明显较小，而微应变较大。所研究的两种蛋白质都没有表现出与IH的特定晶面的优先相互作用。这一结果与药物蛋白与冰的间接相互作用相一致，在冰结晶前沿附近的准液层中蛋白质分子聚集。直接相互作用是指蛋白质分子在冰晶上的吸附，而“间接相互作用”这个术语用来描述蛋白质与冰晶之间的任何干扰，而不涉及吸附。另一方面，溶菌酶分子没有表现出与冰晶(直接或间接)相互作用的任何证据。据我们所知，这是第一份关于不同类型的非抗冻蛋白之间蛋白质/冰相互作用的主要差异的报告。此外，我们还报告了第二组冰晶的意外发现，冰晶结构域的尺寸要小得多(几纳米)。第二个(较小的)群体被初步确定为高压形式的冰，可能是冰III或冰九。这一观察结果突出了机械应力和局部压力在冷冻引起的蛋白质失稳中的潜在作用。
While proteins can often be stabilized by maintaining them in the frozen state, water-to-ice transformation can also lead to degradation of protein molecules. A new method to study protein/ice interaction is presented herein, which is based on measuring the characteristic features of X-ray diffraction (XRD) patterns of hexagonal ice (Ih). Aqueous solutions of four different proteins and several small molecular weight solutes are studied using high-resolution synchrotron X-ray diffraction at the ID22 beamline at the European Synchrotron Radiation Facility. The beamline is optimized to eliminate the instrumental broadening of diffraction lines and reduce the preferred orientation effects, thereby enabling quantitative analysis of the XRD data. The analysis demonstrates that two pharmaceutical proteins, recombinant human albumin (rHA) and monoclonal antibody (mAb), have a pronounced effect on the properties of ice crystals. In particular, the size of the crystalline domains is significantly smaller, and the microstrain is larger, in the solutions of the pharmaceutical proteins, when compared with a model protein (lysozyme), an antifreeze protein, and sucrose and histidine. Neither of the proteins studied exhibit preferred interaction with specific crystalline faces of Ih. The results are consistent with indirect interaction of the pharmaceutical proteins with ice, in which protein molecules are accumulated in the quasi-liquid layer next to growing ice crystallization front. Direct interaction would indicate a sorption of protein molecules on ice crystals, whereas “indirect interaction” terminology is used to describe any interference of proteins with ice crystals without sorption involved. Lysozyme molecules, on the other hand, do not exhibit any evidence of interaction (either direct or indirect) with ice crystals. This is the first report, to the best of our knowledge, of major difference in protein/ice interaction between different types of non-antifreeze proteins. In addition, we report an unexpected finding of a second population of ice crystals, with a much smaller (a few nm) size of crystalline domains. The second (minor) population is tentatively identified as a high-pressure form of ice, possibly IceIII or IceIX. This observation highlights a potential role of mechanical stresses and local pressure in freeze-induced destabilization of proteins.
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DOI：
10.1073/pnas.1212826109
发表时间：
2012-12-04
影响因子：
11.1
作者：
Huang, Mia L.;Ehre, David;Ward, Michael D.
通讯作者：
Ward, Michael D.
DOI：
10.6028/jres.109.005
发表时间：
2004-01-01
影响因子：
1.5
作者：
Kamminga, JD;Seijbel, LJ
通讯作者：
Seijbel, LJ
DOI：
10.1073/pnas.1100429108
发表时间：
2011-05-03
影响因子：
11.1
作者：
Garnham, Christopher P.;Campbell, Robert L.;Davies, Peter L.
通讯作者：
Davies, Peter L.
影响因子：
64.8
作者：
Lobban, C;Finney, JL;Kuhs, WF
通讯作者：
Kuhs, WF
影响因子：
3.4
作者：
BRONSHTEYN, VL;STEPONKUS, PL
通讯作者：
STEPONKUS, PL