Publication: The C-terminal domain of 4-hydroxyphenylacetate 3-hydroxylase from Acinetobacter baumannii is an autoinhibitory domain
Issued Date
2012-07-27
Resource Type
ISSN
1083351X
00219258
00219258
Other identifier(s)
2-s2.0-84864381297
Rights
Mahidol University
Rights Holder(s)
SCOPUS
Bibliographic Citation
Journal of Biological Chemistry. Vol.287, No.31 (2012), 26213-26222
Suggested Citation
Thanawat Phongsak, Jeerus Sucharitakul, Kittisak Thotsaporn, Worrapoj Oonanant, Jirundon Yuvaniyama, Jisnuson Svasti, David P. Ballou, Pimchai Chaiyen The C-terminal domain of 4-hydroxyphenylacetate 3-hydroxylase from Acinetobacter baumannii is an autoinhibitory domain. Journal of Biological Chemistry. Vol.287, No.31 (2012), 26213-26222. doi:10.1074/jbc.M112.354472 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/13659
Research Projects
Organizational Units
Authors
Journal Issue
Thesis
Title
The C-terminal domain of 4-hydroxyphenylacetate 3-hydroxylase from Acinetobacter baumannii is an autoinhibitory domain
Other Contributor(s)
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.