Document Detail


Purification, characterization, gene cloning and nucleotide sequencing of D: -stereospecific amino acid amidase from soil bacterium: Delftia acidovorans.
MedLine Citation:
PMID:  15959727     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The D-amino acid amidase-producing bacterium was isolated from soil samples using an enrichment culture technique in medium broth containing D-phenylalanine amide as a sole source of nitrogen. The strain exhibiting the strongest activity was identified as Delftia acidovorans strain 16. This strain produced intracellular D-amino acid amidase constitutively. The enzyme was purified about 380-fold to homogeneity and its molecular mass was estimated to be about 50 kDa, on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme was active preferentially toward D-amino acid amides rather than their L-counterparts. It exhibited strong amino acid amidase activity toward aromatic amino acid amides including D-phenylalanine amide, D-tryptophan amide and D-tyrosine amide, yet it was not specifically active toward low-molecular-weight D-amino acid amides such as D-alanine amide, L-alanine amide and L-serine amide. Moreover, it was not specifically active toward oligopeptides. The enzyme showed maximum activity at 40 degrees C and pH 8.5 and appeared to be very stable, with 92.5% remaining activity after the reaction was performed at 45 degrees C for 30 min. However, it was mostly inactivated in the presence of phenylmethanesulfonyl fluoride or Cd2+, Ag+, Zn2+, Hg2+ and As3+ . The NH2 terminal and internal amino acid sequences of the enzyme were determined; and the gene was cloned and sequenced. The enzyme gene damA encodes a 466-amino-acid protein (molecular mass 49,860.46 Da); and the deduced amino acid sequence exhibits homology to the D-amino acid amidase from Variovorax paradoxus (67.9% identity), the amidotransferase A subunit from Burkholderia fungorum (50% identity) and other enantioselective amidases.
Authors:
Tipparat Hongpattarakere; Hidenobu Komeda; Yasuhisa Asano
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2005-06-16
Journal Detail:
Title:  Journal of industrial microbiology & biotechnology     Volume:  32     ISSN:  1367-5435     ISO Abbreviation:  J. Ind. Microbiol. Biotechnol.     Publication Date:  2005 Dec 
Date Detail:
Created Date:  2005-12-05     Completed Date:  2006-02-14     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  9705544     Medline TA:  J Ind Microbiol Biotechnol     Country:  England    
Other Details:
Languages:  eng     Pagination:  567-76     Citation Subset:  IM    
Affiliation:
Faculty of Agro-Industry, Prince of Songkla University, Hat Yai, 90112, Songkla, Thailand. tipparat.h@psu.ac.th
Data Bank Information
Bank Name/Acc. No.:
GENBANK/AB154822
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MeSH Terms
Descriptor/Qualifier:
Amidohydrolases* / chemistry,  genetics,  isolation & purification,  metabolism
Amino Acid Sequence
Cloning, Molecular
Delftia acidovorans / enzymology*,  genetics,  isolation & purification
Molecular Sequence Data
Sequence Analysis, DNA
Soil Microbiology*
Stereoisomerism
Substrate Specificity
Chemical
Reg. No./Substance:
EC 3.5.-/Amidohydrolases; EC 3.5.1.-/D-amino acid amidase

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


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