| 项目编号: | 1403077
|
| 项目名称: | SusChEM: Engineering and Evolution of Cytochrome P450 Enzymes for Non-Natural Chemistry |
| 作者: | Frances Arnold
|
| 承担单位: | California Institute of Technology
|
| 批准年: | 2013
|
| 开始日期: | 2014-07-01
|
| 结束日期: | 2017-06-30
|
| 资助金额: | USD345000
|
| 资助来源: | US-NSF
|
| 项目类别: | Standard Grant
|
| 国家: | US
|
| 语种: | 英语
|
| 特色学科分类: | Engineering - Chemical, Bioengineering, Environmental, and Transport Systems
|
| 英文关键词: | bacterial cytochrome p450
; p450 structure
; modern protein engineering technique
; evolutionary potential
; non-natural
; true evolutionary strategy
; known enzyme-catalyzed transformation
; cytochrome p450-bm3
; evolutionary method
; important new enzyme function
; natural enzyme
; evolutionary innovation
; biochemical engineering program
; metabolic engineering
; synthetic chemistry
|
| 英文摘要: | 1403077
Arnold, Frances H.
A future chemicals industry that is sustainable and environmentally friendly will make increasing use of biological processes to convert both renewable and fossil resources to useful chemicals. To date, the use of engineered microbes to produce fuels and chemicals has depended on reassembling existing enzymes into biosynthetic pathways. Unfortunately, many desired products fall outside the reach of the rather limited set of known enzyme-catalyzed transformations or can be made more efficiently using synthetic chemistry. Thus, a future where metabolic engineering might produce nearly all of the organic molecules upon which society depends is still a ways off. Progress in biological production will depend on our ability to genetically encode new catalysts. The goal of this research is to create new cyclopranation catalysts by engineering of a natural enzyme, a bacterial cytochrome P450. Preliminary results indicate that this is possible - the newly-discovered cyclopropanation reaction will be optimized and expanded using modern protein engineering techniques.
Catalytic innovations in nature are rare events and difficult to observe. It is also difficult to discover entirely new activities in the laboratory by evolutionary methods designed to mimic nature. Recently, however, the PI's lab at Caltech used chemical intuition to jumpstart the process and discover that variants of cytochrome P450-BM3 are efficient catalysts of formal carbene transfers when provided with appropriate synthetic reagents. This highly desirable non-natural activity could be increased significantly using semi-rational mutagenesis based on knowledge of the P450 structure and catalytic mechanism. Additional avenues of investigation are needed to continue improving these catalysts and to discover new ones. A three-tiered strategy has been designed to test specific hypotheses regarding improvements while also allowing for evolutionary innovation. Specifically, (i) the effect of heme redox potential on non-natural reactivity will be investigated, with the expectation that achieving more oxidizing redox potentials will increase reactivity; (ii) key conserved residues will be examined by determining the effect of mutations on kinetic parameters and overall turnover levels. (3) a true evolutionary strategy will be employed to optimize non-natural activity using a novel high-throughput cyclopropanation screen and identify and characterize beneficial mutations. This research will both explore and explain the evolutionary potential of this important new enzyme function.
This award is co-funded by the Biotechnology and Biochemical Engineering Program of the CBET Division and by the Systems and Synthetic Biology Program of the Division of Molecular and Cellular Biology. |
| 资源类型: | 项目
|
| 标识符: | http://119.78.100.158/handle/2HF3EXSE/96578
|
| Appears in Collections: | 影响、适应和脆弱性
气候减缓与适应
|
|
There are no files associated with this item.
|
| Recommended Citation: | |
Frances Arnold. SusChEM: Engineering and Evolution of Cytochrome P450 Enzymes for Non-Natural Chemistry. 2013-01-01.
|
|
|