Document Detail


Structure-guided saturation mutagenesis of N-acetylneuraminic acid lyase for the synthesis of sialic acid mimetics.
MedLine Citation:
PMID:  15897188     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Analogues of N-acetylneuraminic acid (sialic acid, NANA, Neu5Ac), including 6-dipropylcarboxamides, have been found to be selective and potent inhibitors of influenza sialidases. Sialic acid analogues are, however, difficult to synthesize by traditional chemical methods and the enzyme N-acetylneuraminic acid lyase (NAL) has previously been used for the synthesis of a number of analogues. The activity of this enzyme towards 6-dipropylcarboxamides is, however, low. Here, we used structure-guided saturation mutagenesis to produce variants of NAL with improved activity and specificity towards 6-dipropylcarboxamides. Three residues were targeted for mutagenesis, Asp191, Glu192 and Ser208. Only substitution at position 192 produced significant improvements in activity towards the dipropylamide. One variant, E192N, showed a 49-fold improvement in catalytic efficiency towards the target analogue and a 690-fold shift in specificity from sialic acid towards the analogue. These engineering efforts provide a scaffold for the further tailoring of NAL for the synthesis of sialic acid mimetics.
Authors:
G J Williams; T Woodhall; A Nelson; A Berry
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2005-05-16
Journal Detail:
Title:  Protein engineering, design & selection : PEDS     Volume:  18     ISSN:  1741-0126     ISO Abbreviation:  Protein Eng. Des. Sel.     Publication Date:  2005 May 
Date Detail:
Created Date:  2005-06-06     Completed Date:  2005-08-30     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  101186484     Medline TA:  Protein Eng Des Sel     Country:  England    
Other Details:
Languages:  eng     Pagination:  239-46     Citation Subset:  IM    
Affiliation:
Astbury Centre for Structural Biology, School of Biochemistry, University of Leeds, Leeds LS2 9JT, UK.
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MeSH Terms
Descriptor/Qualifier:
Amides / metabolism
Amino Acid Substitution
Biomimetics*
Escherichia coli / enzymology*
Molecular Structure
Mutagenesis*
N-Acetylneuraminic Acid / metabolism*
Oxo-Acid-Lyases / chemistry*,  genetics*
Protein Engineering*
Structure-Activity Relationship
Substrate Specificity
Chemical
Reg. No./Substance:
0/Amides; 131-48-6/N-Acetylneuraminic Acid; EC 4.1.3.-/Oxo-Acid-Lyases; EC 4.1.3.3/N-acetylneuraminate lyase

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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