fulltext.study @t Gmail

Enhanced Enzyme Stability Through Site-Directed Covalent Immobilization

Paper ID Volume ID Publish Year Pages File Format Full-Text
22973 43408 2015 8 PDF Available
Title
Enhanced Enzyme Stability Through Site-Directed Covalent Immobilization
Abstract

•We demonstrate a covalent, site-specific enzyme immobilization technology.•The method enables immobilization at specifically chosen sites throughout the enzyme.•Enzyme immobilization orientation significantly affects enzyme activity.•Immobilization orientation control enables greater stability than random attachment.

Breakthroughs in enzyme immobilization have enabled increased enzyme recovery and reusability, leading to significant decreases in the cost of enzyme use and fueling biocatalysis growth. However, current enzyme immobilization techniques suffer from leaching, enzyme stability, and recoverability and reusability issues. Moreover, these techniques lack the ability to control the orientation of the immobilized enzymes. To determine the impact of orientation on covalently immobilized enzyme activity and stability, we apply our PRECISE (Protein Residue-Explicit Covalent Immobilization for Stability Enhancement) system to a model enzyme, T4 lysozyme. The PRECISE system uses non-canonical amino acid incorporation and the Huisgen 1,3-dipolar cycloaddition “click” reaction to enable directed enzyme immobilization at rationally chosen residues throughout an enzyme. Unlike previous site-specific systems, the PRECISE system is a truly covalent immobilization method. Utilizing this system, enzymes immobilized at proximate and distant locations from the active site were tested for activity and stability under denaturing conditions. Our results demonstrate that orientation control of covalently immobilized enzymes can provide activity and stability benefits exceeding that of traditional random covalent immobilization techniques. PRECISE immobilized enzymes were 50 and 73% more active than randomly immobilized enzymes after harsh freeze–thaw and chemical denaturant treatments.

Keywords
Biocatalysis; Enzyme immobilization; Enzyme stability; Non-canonical amino acids; Green manufacturing; Cell-free protein synthesis.
First Page Preview
Enhanced Enzyme Stability Through Site-Directed Covalent Immobilization
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 193, 10 January 2015, Pages 83–90
Authors
, , , , ,
Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us