Publication: Esterase activity of Bordetella pertussis CyaC-acyltransferase against synthetic substrates: Implications for catalytic mechanism in vivo
Issued Date
2010-03-01
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ISSN
15746968
03781097
03781097
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2-s2.0-77149157491
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Mahidol University
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SCOPUS
Bibliographic Citation
FEMS Microbiology Letters. Vol.304, No.2 (2010), 183-190
Suggested Citation
Niramon Thamwiriyasati, Busaba Powthongchin, Jongrak Kittiworakarn, Gerd Katzenmeier, Chanan Angsuthanasombat Esterase activity of Bordetella pertussis CyaC-acyltransferase against synthetic substrates: Implications for catalytic mechanism in vivo. FEMS Microbiology Letters. Vol.304, No.2 (2010), 183-190. doi:10.1111/j.1574-6968.2010.01896.x Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/28771
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Title
Esterase activity of Bordetella pertussis CyaC-acyltransferase against synthetic substrates: Implications for catalytic mechanism in vivo
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Abstract
Adenylate cyclase-hemolysin toxin (CyaA) produced from the human respiratory tract pathogen Bordetella pertussis requires fatty-acyl modification by CyaC-acyltransferase to become an active toxin. Previously, the recombinant CyaA pore-forming (CyaA-PF) fragment expressed in Escherichia coli was shown to be hemolytically active upon palmitoylation in vivo by cosynthesized CyaC. Here, the 21-kDa CyaC enzyme separately expressed in E. coli as an inclusion body was solubilized in 8 M urea and successfully refolded into an enzymatically active monomer. In addition to the capability of activating CyaA-PF in vitro, CyaC showed esterase activity against p-nitrophenyl acetate (pNPA) and p-nitrophenyl palmitate (pNPP), with preferential hydrolysis toward pNPP when compared with chymotrypsin. A homology-based CyaC structure suggested a conceivable role of a catalytic triad including Ser30, His33 and Tyr 66 in substrate catalysis. Alanine substitutions of these individual residues caused a drastic decrease in specific activities of all three mutant enzymes (S30A, H33A and Y66A) toward pNPP, signifying that CyaC-acyltransferase shares a similar mechanism of hydrolysis with a serine esterase in which Ser30 is part of the catalytic triad. © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.