High-yield novel leech hyaluronidase to expedite the preparation of specific hyaluronan oligomers.
High-yield novel leech hyaluronidase to expedite the preparation of specific hyaluronan oligomers.
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高产新型水蛭透明质酸酶可加速特定透明质酸寡聚物的制备
DOI：
10.1038/srep04471
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发表时间：
2014-03-26
影响因子：
4.6
通讯作者：
Chen J
中科院分区：
文献类型：
--
作者：
Jin P;Kang Z;Zhang N;Du G;Chen J
透明质酸酶(Haases)，特别是水蛭透明质酸酶，因其在医学上的广泛应用和在酶法生产透明质酸低聚糖方面的巨大潜力而引起了人们的广泛关注。然而，人们对这第三个有趣的哈斯家族知之甚少。在这里，我们应用随机扩增cDNA末端聚合酶链式反应(RACE-PCR)的方法来鉴定第一个水蚤HAase编码基因。将蛋白质工程与高密度培养相结合，在毕赤酵母分泌型表达系统中实现了−1的高效表达(8.42×105Uml)。与市售牛睾丸HAase相比，重组水蛭HAase具有更好的酶学性质。此外，对水解过程的分析表明，这种新型酶采用非过程性内切模式，产生不同孵育时间的窄谱HA寡糖。该酶的规模化生产不仅将极大地促进医用领域的应用，而且将有利于酶法生产特定的HA寡糖。
Hyaluronidases (HAases), particularly leech HAases, have attracted intense attention due to their broad applications in medical treatments and great potential for the enzymatic production of hyaluronan oligosaccharides. However, little is known about this third interesting family of HAases. Here, we applied the random amplification of cDNA ends polymerase chain reaction (RACE-PCR) approach to identify the first leech HAase-encoding gene. By combining protein engineering and high-density culture, we achieved high-level production (8.42 × 105U ml−1) in the yeastPichia pastorissecretory expression system. Compared with the commercial bovine testicular HAase, the recombinant leech HAase exhibited superior enzymatic properties. Furthermore, analysis of the hydrolytic process suggested that this novel enzyme adopts a nonprocessive endolytic mode, yielding a narrow-spectrum of specific HA oligosaccharides with different incubation times. Large-scale production of this novel leech HAase will not only greatly promote medical applications but also facilitate the enzymatic production of specific HA oligosaccharides.
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影响因子：
6.4
作者：
Liu L;Liu Y;Li J;Du G;Chen J
通讯作者：
Chen J
影响因子：
11.2
作者：
Benitez A;Yates TJ;Lopez LE;Cerwinka WH;Bakkar A;Lokeshwar VB
通讯作者：
Lokeshwar VB
影响因子：
4.3
作者：
Mahoney, DJ;Aplin, RT;Day, AJ
通讯作者：
Day, AJ
影响因子：
4.8
作者：
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通讯作者：
Kaneko, Satoshi
影响因子：
46.9
作者：
De Schutter, Kristof;Lin, Yao-Cheng;Callewaert, Nico
通讯作者：
Callewaert, Nico