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Improved expression and characterization of recombinant human Golgi α1,2-mannosidase I isoforms (IA2 and IC) by Escherichia coli

Paper ID Volume ID Publish Year Pages File Format Full-Text
21179 43210 2011 6 PDF Available
Title
Improved expression and characterization of recombinant human Golgi α1,2-mannosidase I isoforms (IA2 and IC) by Escherichia coli
Abstract

Golgi α1,2-mannosidase I is involved in the N-linked oligosaccharide processing pathway. In this study, two truncated genes encoding for human Golgi α1,2-mannosidase I (hManIA2: amino acids 127–626 and hManIC: amino acids 118–617) were expressed in Escherichia coli to characterize the enzymes. These genes were fused to a T7 protein tag and a histidine tag at the N- and C-terminal ends, respectively, and purified using Co2+ affinity chromatography. The properties including optimal temperature, optimal pH, and substrate specificity of the purified enzymes were investigated by HPLC using pyridylamino (PA)-labeled oligosaccharides as substrates. The stability of hManIA2 was dependent on the presence of Ca2+, which was also required for its activity. On the other hand, hManIC was stable in the absence of Ca2+, even though Ca2+ was also effective for the activity of hManIC. While the similarity of the amino acid sequences is over 60%, hManIA2 and hManIC showed different substrate specificities particularly toward M9A and M8C.

Keywords
Human α1,2-mannosidase I; Non-inductive expression; Escherichia coli; N-glycan
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Improved expression and characterization of recombinant human Golgi α1,2-mannosidase I isoforms (IA2 and IC) by Escherichia coli
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 112, Issue 1, July 2011, Pages 14–19
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