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N-Acetyl-d-glucosamine 2-epimerase from Anabaena sp. CH1 contains a novel ATP-binding site required for catalytic activity

Paper ID Volume ID Publish Year Pages File Format Full-Text
35149 45078 2012 5 PDF Available
Title
N-Acetyl-d-glucosamine 2-epimerase from Anabaena sp. CH1 contains a novel ATP-binding site required for catalytic activity
Abstract

ATP is required as a structural activator for the reversible epimerization of N-acetyl-d-glucosamine to N-acetyl-d-mannosamine by N-acetyl-d-glucosamine 2-epimerase (AGE); however, the ATP-binding site on AGE has not been clearly identified. This study aimed to investigate the specific region of Anabaena sp. CH1 AGE (bAGE) that is required for ATP binding. In the absence of ATP, tryptic digest of bAGE resulted in the production of 2 segments of 17 and 26 kDa, while in the presence of 1 mM ATP, the enzyme was resistant to trypsin. ADP also displayed protective effects against trypsin digestion. A trypsin-mediated ATP-footprinting assay identified a deviant ATP-protected region, 156-GKYTK-160, which is located within the flexible loop of bAGE. Site-directed mutagenesis of residues in the loop region was performed, and both K151A and K160A variants greatly decreased the enzymatic activity as well as the ATP-binding ability of bAGE, indicating that residues K151 and K160 may be critical for ATP binding. This study demonstrated that the ATP-binding site (151-KDNPKGKYTK-160) of bAGE was a novel rather than a classical Walker motif A. This is the first ATP-binding site reported for AGEs.

► We study the ATP binding site of Anabaena sp. CH1 GlcNAc 2-epimerase (bAGE). ► bAGE has an ATP-footprinting property. ► ATP binding site is located in a flexible loop with lysine, arginine and glycine. ► Residues K151 and K160 of bAGE might be critical for ATP binding. ► The ATP binding region of bAGE is novel, rather than a classical Walker motif A.

Keywords
N-Acetyl-d-glucosamine 2-epimerase; Anabaena sp.; ATP-binding site; Walker motif A
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N-Acetyl-d-glucosamine 2-epimerase from Anabaena sp. CH1 contains a novel ATP-binding site required for catalytic activity
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 47, Issue 6, June 2012, Pages 948–952
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