The 0.78 Å structure of a serine protease:: Bacillus lentus subtilisin
The 0.78 Å structure of a serine protease:: Bacillus lentus subtilisin
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DOI：
10.1021/bi9813983
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发表时间：
1998-09-29
期刊：
影响因子：
2.9
通讯作者：
Bott, R
中科院分区：
文献类型：
--
作者：
Kuhn, P;Knapp, M;Bott, R
超高分辨率X射线衍射数据从低温冷却，B。已收集到分辨率为0.78埃的迟缓菌枯草杆菌蛋白酶晶体。相对于在室温和2.0埃分辨率下测定的相同结构，细化的模型坐标具有0.22埃的rms偏差。在一条多肽链的269个残基中，有21个残基被鉴定为主链和侧链紊乱的几个区域。氢原子在F-o - F-c差分电子密度图中显示为显著峰，并且可以区分碳、氮和氧原子。所有主链非氢键长度的估计标准偏差（ESD）为0.009埃，键角为0.5度，基于无限制的全矩阵最小二乘改进。氢键在丝氨酸蛋白酶催化三联体（Ser-His-Asp）中被分解。电子密度观察到一个不寻常的，短的氢键之间的天冬氨酸和组氨酸的催化三联体。在许多丝氨酸蛋白酶中通过NMR鉴定的氢原子似乎由键中的杂原子共享。这是第一次报道的详细的化学特征之间的相关性，通过NMR和那些在低温冷却的晶体结构测定在1000分辨率。短的氢键，称为“催化氢键”，作为一个复杂的氢键网络的一部分，涉及催化三元组的天冬氨酸。虽然不寻常，这些功能似乎有保守的类似物在其他丝氨酸蛋白酶家族，虽然具体细节不同的家庭。
Ultrahigh-resolution X-ray diffraction data from cryo-cooled, B. lentus subtilisin crystals has been collected to a resolution of 0.78 Angstrom. The refined model coordinates have a rms deviation of 0.22 Angstrom relative to the same structure determined at room temperature and 2.0 Angstrom resolution. Several regions of main-chain and side-chain disorder have been identified for 21 our of 269 residues in one polypeptide chain. Hydrogen atoms appear as significant peaks in the F-o - F-c difference electron density map, and carbon, nitrogen, and oxygen atoms can be differentiated. The estimated standard deviation (ESD) for all main-chain non-hydrogen bond lengths is 0.009 Angstrom and 0.5 degrees for bond angles based on an unrestrained full-matrix least-squares refinement. Hydrogen bonds are resolved in the serine protease catalytic triad (Ser-His-Asp). Electron density is observed for an unusual, short hydrogen bond between aspartic acid and histidine in the catalytic triad. The hydrogen atom, identified by NMR in numerous serine proteases, appears to be shared by the heteroatoms in the bond. This represents the first reported correlation between detailed chemical features identified by NMR and those in a cryo-cooled crystallographic structure determination at ultrahigh resolution. The short hydrogen bond, designated "catalytic hydrogen bond", occurs as part of an elaborate hydrogen bond network, involving Asp of the catalytic triad. While unusual, these features appear to have conserved analogues in other serine protease families although specific details differ from family to family.