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Functional immobilization of recombinant alkaline phosphatases bearing a glutamyl donor substrate peptide of microbial transglutaminase

Paper ID Volume ID Publish Year Pages File Format Full-Text
22497 43295 2007 5 PDF Available
Title
Functional immobilization of recombinant alkaline phosphatases bearing a glutamyl donor substrate peptide of microbial transglutaminase
Abstract

Covalent and site-specific protein immobilization catalyzed by microbial transglutaminase (MTG) was investigated using recombinant Escherichia coli alkaline phosphatase (AP) tagged with a glutamyl donor substrate peptide (MLAQGS) of MTG. A polystyrene surface physically coated with β-casein or bovine serum albumin (BSA) was employed as an MTG-specific surface displaying reactive lysine residues. MTG-mediated protein immobilization through catalytic ɛ-(γ-glutamyl)lysine bond formation between the peptide tag of recombinant APs and β-casein- or BSA-coated surface was verified by the detection of AP activity on the surface. It was found that the length and the insertion position of the peptide tag did not significantly affect the efficacy of enzymatic immobilization of the recombinant APs. On the other hand, pH and ionic strength in the reaction media had crucial effects on the immobilization yields. Interestingly, the optimum pH range of MTG-mediated protein immobilization differed markedly from that for an MTG-catalyzed reaction in aqueous solution. The results suggest that the concentration of reactive species due to electrostatic interaction between the enzyme-substrate intermediate and the protein-adsorbed surface is a key factor governing MTG catalysis at a solid surface.

Keywords
enzymatic immobilization; peptide tag; posttranslational modification; site-specific modification; transglutaminase
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Functional immobilization of recombinant alkaline phosphatases bearing a glutamyl donor substrate peptide of microbial transglutaminase
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 104, Issue 3, September 2007, Pages 195–199
Authors
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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