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Please use this identifier to cite or link to this item: http://repository.li.mahidol.ac.th/dspace/handle/123456789/13659
Title: The C-terminal domain of 4-hydroxyphenylacetate 3-hydroxylase from Acinetobacter baumannii is an autoinhibitory domain
Authors: Thanawat Phongsak
Jeerus Sucharitakul
Kittisak Thotsaporn
Worrapoj Oonanant
Jirundon Yuvaniyama
Jisnuson Svasti
David P. Ballou
Pimchai Chaiyen
Mahidol University
Chulalongkorn University
University of Michigan, Ann Arbor
Keywords: Biochemistry, Genetics and Molecular Biology
Issue Date: 27-Jul-2012
Citation: Journal of Biological Chemistry. Vol.287, No.31 (2012), 26213-26222
Abstract: p-Hydroxyphenylacetate (HPA) 3-hydroxylase from Acinetobacter baumannii consists of a reductase component (C 1 ) and an oxygenase component (C 2 ). C 1 catalyzes the reduction of FMN by NADH to provide FMNH - as a substrate for C 2 . The rate of reduction of flavin is enhanced ∼20-fold by binding HPA. The N-terminal domain of C1 is homologous to other flavin reductases, whereas the C-terminal domain (residues 192-315) is similar to MarR, a repressor protein involved in bacterial antibiotic resistance. In this study, three forms of truncated C 1 variants and single site mutation variants of residues Arg-21, Phe-216, Arg-217, Ile-246, and Arg-247 were constructed to investigate the role of the C-terminal domain in regulating C 1 . In the absence of HPA, the C 1 variant in which residues 179-315 were removed (t178C 1 ) was reduced by NADH and released FMNH - at the same rates as wild-type enzyme carries out these functions in the presence of HPA. In contrast, variants with residues 231-315 removed behaved similarly to the wild-type enzyme. Thus, residues 179-230 are involved in repressing the production of FMNH - in the absence of HPA. These results are consistent with the C-terminal domain in the wild-type enzyme being an autoinhibitory domain that upon binding the effector HPA undergoes conformational changes to allow faster flavin reduction and release. Most of the single site variants investigated had catalytic properties similar to those of the wild-type enzyme except for the F216A variant, which had a rate of reduction that was not stimulated by HPA. F216A could be involved with HPA binding or i n the required conformational change for stimulation of flavin reduction by HPA. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.
URI: https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84864381297&origin=inward
http://repository.li.mahidol.ac.th/dspace/handle/123456789/13659
ISSN: 1083351X
00219258
Appears in Collections:Scopus 2011-2015

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