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Engineering an Enzyme for Direct Electrical Monitoring of Activity.

ABSTRACT: Proteins have been shown to be electrically conductive if tethered to an electrode by means of a specific binding agent, allowing single molecules to be wired into an electrical sensing circuit. Such circuits allow enzymes to be used as sensors, detectors, and sequencing devices. We have engineered contact points into a ?29 polymerase by introducing biotinylatable peptide sequences. The modified enzyme was bound to electrodes functionalized with streptavidin. ?29 connected by one biotinylated contact, and a second nonspecific contact showed rapid small fluctuations in current when activated. Signals were greatly enhanced with two specific contacts. Features in the distributions of DC conductance increased by a factor 2 or more over the open to closed conformational transition of the polymerase. Polymerase activity is manifested by a rapid (millisecond) large (25% of background) current fluctuations imposed on the DC conductance.

SUBMITTER: Zhang B

PROVIDER: S-EPMC7047563 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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Engineering an Enzyme for Direct Electrical Monitoring of Activity.

Zhang Bintian B Deng Hanqing H Mukherjee Sohini S Song Weisi W Wang Xu X Lindsay Stuart S

ACS nano 20191015 2

Proteins have been shown to be electrically conductive if tethered to an electrode by means of a specific binding agent, allowing single molecules to be wired into an electrical sensing circuit. Such circuits allow enzymes to be used as sensors, detectors, and sequencing devices. We have engineered contact points into a Φ29 polymerase by introducing biotinylatable peptide sequences. The modified enzyme was bound to electrodes functionalized with streptavidin. Φ29 connected by one biotinylated co ...[more]

PMID: 31594304

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Project description:The article investigates electrically active ceramic composite of [Formula: see text] (HAP) and [Formula: see text] (BST) for biomedical applications. The study is a systematic blend of the materials science aspect of composites with a special focus on the dielectric and biological properties and their relationships. The article emphasized primarily extracting the dielectric constant ([Formula: see text] of the specimens (that lay in the range of 3-65) and related them to microstructural properties like the grain size and at.% of BST. A broad outlook on the importance of [Formula: see text] in determining the suitability of bioceramics for clinical applications is presented. Bioactivity analysis of the specimens led to probing the surface charges (that were negative), and it was found crucial to the growth of dense apatite layers. Furthermore, the cytocompatibility of the specimens displayed cell viability above 100% for Day 1, which increased substantially for Day 3. To reveal other biological properties of the composites, protein adsorption studies using bovine serum albumin (BSA) and fetal bovine serum (FBS) was carried out. Electrostatic interactions govern the adsorption, and the mathematical dependence on surface charges is linear. The protein adsorption is also linearly correlated with the [Formula: see text], intrinsic to the biomaterials. We delve deeper into protein-biomaterials interactions by considering the evolution of the secondary structure of BSA adsorbed into the specimens. Based on the investigations, 20 at.% HAP-80 at.% BST (20H-80B) was established as a suitable composite comprising the desired features of HAP and BST. Such explorations of electrical and biological properties are interesting for modulating the behavior of bioceramic composites. The results project the suitability of 20H-80B for designing electrically active smart scaffolds for the proposed biomedical applications and are expected to incite further clinical trials.

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Engineering an Enzyme for Direct Electrical Monitoring of Activity.

Publications

Engineering an Enzyme for Direct Electrical Monitoring of Activity.

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