One of the key fundamental scientific questions is how isolated enzymes maintain their native active conformations in solution or in immobilization matrix. Our long-term goal is to elucidate the mechanisms for enzyme activity enhancements in functionalized nanoporous support to exploit highly-active and stable enzymes for detoxification, cancer treatment, biosensing, protein drug release and delivery. The specific hypothesis is that: a protein's enzymatic activity and stability can be significantly enhanced in an appropriately engineered open nanoporous support, which functions as a confined and interactive nanoenvironment for promoting a favorable protein conformational change. This hypothesis is based on the observations: First, we have entrapped three different enzymes in functionalized mesoporous silica (FMS). Mesoporous silica is a typical open nanoporous support with pore sizes as large as tens of nanometers. We demonstrated that all the three enzymes exhibit enhanced activity in FMS in comparison with the enzymes free in solution;Second, enzyme-specific activity can be increased or decreased to a large extent by changing protein loading density in FMS;Third, we found that FMS and chaotropic agents can act synergistically to enhance enzyme activity;Fourth, we found experimental evidences indicating there were favorable protein conformational changes occurring in FMS. We believe that, (i) FMS is a confined space, and (ii) FMS provides an interactive environment promoting a favorable protein conformational change, thereby enhancing enzyme activity and stability. Therefore, we propose the specific aims to: 1. Investigate necessity of mesoporous structure and effects of mesopore sizes on the enzyme activity enhancement;2. Investigate the interactions of proteins with FMS to understanding FMS confinement and interactive effects on enzyme activity enhancement;3. Develop molecular models and employ molecular docking and molecular dynamics simulations to probe the mechanism by which FMS steers enzyme conformational dynamics towards enhanced activity;4. Evaluate the efficacy of highly-active and stable organophosphorus hydrolase in FMS to provide the in vivo detoxification towards organophosphorus neurotoxicity in the rat, to demonstrate an integrated all-in-one device of protein (enzyme) drug storage, release, and delivery. Project narrative One of the key fundamental scientific questions is how isolated enzymes maintain their native active conformations in solution or in immobilization matrix. Our long-term goal is to elucidate the mechanisms for enzyme activity enhancements in engineered nanoporous support to exploit highly-active and stable enzymes for medical applications including diagnostics, detoxification, and treatment for cancer and other diseases. As a result of this effort, we will evaluate the efficacy of highly-active and stable organophosphorus hydrolase in the functional nanoporous support to provide the in vivo detoxification towards organophosphorus neurotoxicity in the rat, to demonstrate an integrated all-in-one device of protein (enzyme) drug storage, release, and delivery.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM080987-05
Application #
8310034
Study Section
Special Emphasis Panel (ZRG1-NANO-M (01))
Program Officer
Lewis, Catherine D
Project Start
2008-08-01
Project End
2014-07-31
Budget Start
2012-08-01
Budget End
2014-07-31
Support Year
5
Fiscal Year
2012
Total Cost
$321,757
Indirect Cost
$125,737
Name
Battelle Pacific Northwest Laboratories
Department
Type
DUNS #
032987476
City
Richland
State
WA
Country
United States
Zip Code
99352
Chen, Baowei; Qi, Wen; Li, Xiaolin et al. (2013) Heated proteins are still active in a functionalized nanoporous support. Small 9:2228-32
Lei, Chenghong; Chen, Baowei; Li, Xiaolin et al. (2013) Non-destructively shattered mesoporous silica for protein drug delivery. Microporous Mesoporous Mater 175:157-160
Chen, Baowei; Shah, Saumil S; Shin, Yongsoon et al. (2012) In vitro release of organophosphorus acid anhydrolase from functionalized mesoporous silica against nerve agents. Anal Biochem 421:477-81
Yu, Yuehua; Chen, Baowei; Qi, Wen et al. (2012) Enzymatic conversion of CO(2) to bicarbonate in functionalized mesoporous silica. Microporous Mesoporous Mater 153:166-170
Tam, Tsz Kin; Chen, Baowei; Lei, Chenghong et al. (2012) In situ regeneration of NADH via lipoamide dehydrogenase-catalyzed electron transfer reaction evidenced by spectroelectrochemistry. Bioelectrochemistry 86:92-6
Gomes, Diego E B; Lins, Roberto D; Pascutti, Pedro G et al. (2011) Conformational variability of organophosphorus hydrolase upon soman and paraoxon binding. J Phys Chem B 115:15389-98
Lei, Chenghong; Liu, Pu; Chen, Baowei et al. (2010) Local release of highly loaded antibodies from functionalized nanoporous support for cancer immunotherapy. J Am Chem Soc 132:6906-7
Gomes, Diego E B; Lins, Roberto D; Pascutti, Pedro G et al. (2010) The role of nonbonded interactions in the conformational dynamics of organophosphorous hydrolase adsorbed onto functionalized mesoporous silica surfaces. J Phys Chem B 114:531-40
Chen, Baowei; Lei, Chenghong; Shin, Yongsoon et al. (2009) Probing mechanisms for enzymatic activity enhancement of organophosphorus hydrolase in functionalized mesoporous silica. Biochem Biophys Res Commun 390:1177-81