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【bio-news】创造人造P450酶
Volume 84, Number 16
p. 7
Computational Biology
Artificial P450 Enzymes Created
Computational method leads to library of enzymes that fold and function
创造人造P450酶
计算技术创造具有折叠功能的酶库
Celia Arnaud
A family of nearly 3,000 artificial cytochrome P450 enzymes has been created by a California Institute of Technology team's efforts to recombine sections of three natural cytochrome P450s, a large family of oxidative enzymes that are widespread in nature (PLoS Biol., published online April 11, dx.doi.org/10.1371/journal.pbio.0040112). In humans, P450s play a crucial role in the metabolism of drugs and other toxins.
加利福尼亚技术研究所小组通过重组三种天然的细胞色素P450酶的片段创造一个具有近3000个人造细胞色素P450酶家族，细胞色素P450酶是在自然界广泛存在的氧化酶大家族(PLoS Biol., published online April 11, dx.doi.org/10.1371/journal.pbio.0040112)。在人类，P450在药物和其他毒素的新陈代谢中起到重要作用。
"I'm hoping that this new family will contain cytochrome P450s that people would want to use, for example, to make the human metabolites of drugs or to synthesize complex, biologically active compounds," says Frances H. Arnold, a chemical engineering professor at Caltech. "It's my dream to make a whole library of cytochrome P450s that could hydroxylate anything."
“我希望这个新家族将包括人类需要的细胞色素P450酶，例如，处理药物新陈代谢或合成复杂的，具生物活性的化合物，”加利福尼亚理工学院的化学工程教授Frances H. Arnold说，“我的梦想就是创造一整个可以羟基化任何物质的细胞色素P450酶库。
Arnold, graduate student Christopher R. Otey, and coworkers used a computational method called SCHEMA to guide the creation of the new protein sequences and increase the likelihood that they will be useful.
Arnold，研究生Christopher R. Otey，还有合作者使用一种称为SCHEMA的计算技术去指导新蛋白序列的构造，增加它们有用的可能性。
P450 enzymes are such a diverse family of enzymes that the usual method of random DNA shuffling would generate a set of new sequences in which most of the members would not fold, Arnold says. "You'd be screening garbage."
P450酶是一个多变的家族，普通的随机DNA混合产生的一套新的序列，其中绝大部分无法折叠，Arnold说“你将在垃圾中不停筛选。”
To generate P450 enzymes that catalyze new reactions, Arnold's team wanted the new proteins to be 70 to 100 amino acids different from the starting proteins, yet still fold properly. They made new proteins by recombining chunks of the P450 enzymes nature has provided, Arnold says. "It's a dual optimization problem. We recombine them to preserve as many structural interactions as possible, while at the same time making lots of mutations."
为了得到催化新的反应P450酶，Arnold的团队需要那些有70-100个氨基酸与原先的蛋白不同的新蛋白，但仍能正常地折叠。他们通过重组自然界中已有的P450酶的大部分获得新蛋白，Arnold说“只是一个双重最优化难题。我们重组他们以保护近可能多的结构间交互作用，同时制造大量突变。
That's where SCHEMA comes in. SCHEMA's job is to improve the likelihood that a given sequence will fold by considering the structures of the parent proteins. The crystal structures of the parent proteins are encoded mathematically to make counting broken interactions between side chains a simple calculation. "SCHEMA penalizes you for every broken contact. It says, 'Thou shalt make a library such that most of the members have few broken interactions,' " Arnold says. "At the end, you get a design that penalizes you the least."
这时需要SCHEMA参与，SCHEMA的工作是通过考虑父本蛋白的结构，改善假定序列折叠的可能性。父本蛋白的晶体结构被数学编码以计算侧链间遭到破坏的交互作用。“SCHEMA告诉你每一个被破坏的连接，就是说，‘你可以创建一个大部分由交互作用破坏得非常少的成员组成的库。’” Arnold说“最后，你得到一个损害最小的设计。”
The Caltech team chopped each of the three original enzymes into eight pieces and recombined them, yielding 6,561 (38) possible sequences. Of those, nearly half fold into properly functioning cytochrome P450s that can catalyze a reaction.
加州理工学院小组把3种酶每个分成8块，然后重组他们，得到6561（38）种可能的序列。其中近一半的可以折叠成有功能能够催化反应的细胞色素P450。
But the nonfolding proteins serve a useful purpose, too. The team used a mathematical technique known as logistic regression analysis, which relies on having sequences that don't fold and function in addition to sequences that do, to glean information about why particular sequences fold. "If you're trying to understand what it is about a sequence of amino acids that makes it into a functional protein, it's nice to have ones that weren't successful with which to compare," Arnold says.
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作者:admin@医学,生命科学 2010-12-12 17:11
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