Laboratory for Conservation and Utilization of Bioresources, Yunnan University, 2 North Cuihu Road, Kunming 650091, China.
FASEB J. 2010 May;24(5):1391-400. doi: 10.1096/fj.09-136408. Epub 2009 Dec 9.
Cuticle-degrading proteases are involved in the breakdown of cuticle/eggshells of nematodes or insects, a hard physical barrier against fungal infections. Understanding the 3-dimensional structures of these proteins can provide crucial information for improving the effectiveness of these fungi in biocontrol applications, e.g., by targeted protein engineering. However, the structures of these proteases remain unknown. Here, we report the structures of two cuticle-degrading proteases from two species of nematophagous fungi. The two structures were solved with X-ray crystallography to resolutions of 1.65 A (Ver112) and 2.1 A (PL646), respectively. Crystal structures of PL646 and Ver112 were found to be very similar to each other, and similar to that of proteinase K from another fungus Tritirachium album. Differences between the structures were found among residues of the substrate binding sites (S1 and S4). Experimental studies showed that the enzymes differed in hydrolytic activity to synthetic peptide substrates. Our analyses of the hydrophobic/hydrophilic and electrostatic features of these two proteins suggest that their surfaces likely play important roles during fungal infection against nematodes. The two crystal structures provide a solid basis for investigating the relationship between structure and function of cuticle-degrading proteases.
角质层降解蛋白酶参与线虫或昆虫的角质层/蛋壳的分解，这是真菌感染的坚硬物理屏障。了解这些蛋白质的三维结构可以为提高这些真菌在生物防治中的有效性提供关键信息，例如通过靶向蛋白质工程。然而，这些蛋白酶的结构仍然未知。在这里，我们报告了两种食线虫真菌的两种角质层降解蛋白酶的结构。这两个结构通过 X 射线晶体学分别以 1.65Å（Ver112）和 2.1Å（PL646）的分辨率得到解决。PL646 和 Ver112 的晶体结构彼此非常相似，也与来自另一种真菌 Tritirachium album 的蛋白酶 K 的结构相似。结构之间的差异存在于底物结合位点（S1 和 S4）的残基中。实验研究表明，这些酶在水解合成肽底物的活性上存在差异。我们对这两种蛋白质的疏水性/亲水性和静电特性的分析表明，它们的表面在真菌对线虫的感染过程中可能发挥重要作用。这两个晶体结构为研究角质层降解蛋白酶的结构与功能关系提供了坚实的基础。
