Hu S H, Parker M W, Lei J Y, Wilce M C, Benian G M, Kemp B E
St. Vincent's Institute of Medical Research, Fitzroy, Victoria, Australia.
Nature. 1994 Jun 16;369(6481):581-4. doi: 10.1038/369581a0.
Many protein kinases are self-regulated by an intrasteric mechanism where part of the enzyme's structure directly inhibits the active site. This inhibitory structure is called a pseudosubstrate and specific regulators are required to remove it from the active site to allow substrates access. Removal of the pseudosubstrate sequence from members of the myosin light-chain kinase subfamily, including twitchin kinase, activates them but it is not known whether the pseudosubstrate sequence binds to the active site. Native twitchin is a 753K protein (6,839 residues) located in muscle A-bands of the nematode Caenorhabditis elegans and because of its size has not been easy to study. We have determined the crystal structure, refined to 2.8 A resolution, of a recombinant fragment (residues 5,890 to 6,262) of twitchin kinase that contains the catalytic core and a 60 residue carboxy-terminal tail. The C-terminal tail extends through the active site, wedged between the small and large lobes of the structure and making extensive contacts with the catalytic core which accounts for autoinhibition and provides direct support for the intrasteric mechanism of protein kinase regulation.
许多蛋白激酶通过一种分子内机制进行自我调节，即酶结构的一部分直接抑制活性位点。这种抑制性结构被称为假底物，需要特定的调节因子将其从活性位点移除，以使底物能够接近。从肌球蛋白轻链激酶亚家族成员（包括抽动蛋白激酶）中去除假底物序列会激活它们，但尚不清楚假底物序列是否与活性位点结合。天然的抽动蛋白是一种753K的蛋白质（6839个残基），位于线虫秀丽隐杆线虫的肌肉A带中，由于其大小，一直难以研究。我们已经确定了抽动蛋白激酶重组片段（残基5890至6262）的晶体结构，该片段包含催化核心和一个60个残基的羧基末端尾巴，结构精修至2.8埃分辨率。羧基末端尾巴穿过活性位点，楔入结构的小结构域和大结构域之间，并与催化核心广泛接触，这解释了自抑制现象，并为蛋白激酶调节的分子内机制提供了直接支持。
