Degradation of tropoelastin by matrix metalloproteinases - cleavage site specificities and release of matrikines
Degradation of tropoelastin by matrix metalloproteinases - cleavage site specificities and release of matrikines
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DOI：
10.1111/j.1742-4658.2010.07616.x
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发表时间：
2010-04-01
期刊：
影响因子：
5.4
通讯作者：
Schmelzer, Christian E. H.
中科院分区：
文献类型：
--
作者：
Heinz, Andrea;Jung, Michael C.;Schmelzer, Christian E. H.
为了为开发治疗弹性蛋白降解性疾病的方法提供基础，本研究的目的是研究弹性蛋白降解基质金属蛋白酶MMP-7、MMP-9和MMP-12对天然底物对抗性弹性蛋白的降解作用，并利用互补质谱技术和分子模型比较这些酶的裂解位点特异性。此外，还研究了这三种蛋白酶释放生物活性肽的能力。Tropoelastin很容易被所有三种MMPs降解。MMP-12在对抗性弹性蛋白中鉴定出89个切割位点，而MMP-7和MMP-9分别仅鉴定出58个和63个切割位点。剪切主要发生在对弹力蛋白的n端和c端区域。关于切割位点的特异性，研究表明，所有三种MMPs在P-1'上都同样耐受疏水和/或脂肪氨基酸，包括Pro、Gly、Ile和Val。MMP-7对P-1′上的Leu表现出强烈的偏好，MMP-9和MMP-12也接受这种偏好。在所有三种MMPs中，MMP-12最能耐受P-1'上的大量带电氨基酸和芳香氨基酸。所有三种MMPs都表现出对Pro at P-3的明显偏好，这可以通过分子模型在结构上解释。对生成的肽的分析显示，所有三种MMPs都显示出相似的释放生物活性序列的能力，其中MMP-12产生的这些肽数量最多。此外，所生成的多肽YTTGKLPYGYGPGG、YGARPGVGVGGIP和PGFGAVPGA含有尚未被证明具有生物活性的GxxPG基序，经生物活性测试被鉴定为新的基质因子。
To provide a basis for the development of approaches to treat elastin-degrading diseases, the aim of this study was to investigate the degradation of the natural substrate tropoelastin by the elastinolytic matrix metalloproteinases MMP-7, MMP-9, and MMP-12 and to compare the cleavage site specificities of the enzymes using complementary MS techniques and molecular modeling. Furthermore, the ability of the three proteases to release bioactive peptides was studied. Tropoelastin was readily degraded by all three MMPs. Eighty-nine cleavage sites in tropoelastin were identified for MMP-12, whereas MMP-7 and MMP-9 were found to cleave at only 58 and 63 sites, respectively. Cleavages occurred predominantly in the N-terminal and C-terminal regions of tropoelastin. With respect to the cleavage site specificities, the study revealed that all three MMPs similarly tolerate hydrophobic and/or aliphatic amino acids, including Pro, Gly, Ile, and Val, at P-1'. MMP-7 shows a strong preference for Leu at P-1', which is also well accepted by MMP-9 and MMP-12. Of all three MMPs, MMP-12 best tolerates bulky charged and aromatic amino acids at P-1'. All three MMPs showed a clear preference for Pro at P-3 that could be structurally explained by molecular modeling. Analysis of the generated peptides revealed that all three MMPs show a similar ability to release bioactive sequences, with MMP-12 producing the highest number of these peptides. Furthermore, the generated peptides YTTGKLPYGYGPGG, YGARPGVGVGGIP, and PGFGAVPGA, containing GxxPG motifs that have not yet been proven to be bioactive, were identified as new matrikines upon biological activity testing.