Document Detail

Chemoenzymatic synthesis of CMP-sialic acid derivatives by a one-pot two-enzyme system: comparison of substrate flexibility of three microbial CMP-sialic acid synthetases.
MedLine Citation:
PMID:  15556760     Owner:  NLM     Status:  MEDLINE    
Three C terminal His6-tagged recombinant microbial CMP-sialic acid synthetases [EC] cloned from Neisseria meningitidis group B, Streptococcus agalactiae serotype V, and Escherichia coli K1, respectively, were evaluated for their ability in the synthesis of CMP-sialic acid derivatives in a one-pot two-enzyme system. In this system, N-acetylmannosamine or mannose analogs were condensed with pyruvate, catalyzed by a recombinant sialic acid aldolase [EC] cloned from E. coli K12 to provide sialic acid analogs as substrates for the CMP-sialic acid synthetases. The substrate flexibility and the reaction efficiency of the three recombinant CMP-sialic acid synthetases were compared, first by qualitative screening using thin layer chromatography, and then by quantitative analysis using high performance liquid chromatography. The N. meningitidis synthetase was shown to have the highest expression level, the most flexible substrate specificity, and the highest catalytic efficiency among the three synthetases. Finally, eight sugar nucleotides, including cytidine 5'-monophosphate N-acetylneuraminic acid (CMP-Neu5Ac) and its derivatives with substitutions at carbon-5, carbon-8, or carbon-9 of Neu5Ac, were synthesized in a preparative (100-200 mg) scale from their 5- or 6-carbon sugar precursors using the N. meningitidis synthetase and the aldolase.
Hai Yu; Hui Yu; Rebekah Karpel; Xi Chen
Publication Detail:
Type:  Comparative Study; Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Bioorganic & medicinal chemistry     Volume:  12     ISSN:  0968-0896     ISO Abbreviation:  Bioorg. Med. Chem.     Publication Date:  2004 Dec 
Date Detail:
Created Date:  2004-11-23     Completed Date:  2005-05-19     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  9413298     Medline TA:  Bioorg Med Chem     Country:  England    
Other Details:
Languages:  eng     Pagination:  6427-35     Citation Subset:  IM    
Department of Chemistry, University of California, One Shields Avenue, Davis, CA 95616, USA.
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MeSH Terms
Cloning, Molecular
Cytidine Monophosphate N-Acetylneuraminic Acid / analogs & derivatives,  chemical synthesis*
Escherichia coli / genetics
N-Acylneuraminate Cytidylyltransferase / metabolism*
Neisseria meningitidis, Serogroup B / genetics
Pyruvic Acid
Recombinant Proteins / chemistry
Streptococcus agalactiae / genetics
Substrate Specificity
Reg. No./Substance:
0/Recombinant Proteins; 127-17-3/Pyruvic Acid; 3063-71-6/Cytidine Monophosphate N-Acetylneuraminic Acid; 31103-86-3/Mannose; EC Cytidylyltransferase

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