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Amino acid substitutions in naphthalene dioxygenase from Pseudomonas sp. strain NCIB 9816-4 result in regio- and stereo-specific hydroxylation of flavanone and isoflavanone.
Appl Microbiol Biotechnol. 2013 Jan; 97(2):693-704.AM

Abstract

Wild-type naphthalene dioxygenase (NDO) from Pseudomonas sp. strain NCIB 9816-4 transforms relatively planar flavone and isoflavone to cis-dihydrodiols. However, this enzyme cannot catalyze the transformation of flavanone and isoflavanone in which a phenyl group bonds to the stereogenic C2 or C3 of the C-ring. Protein modeling suggested that Phe224 in the substrate binding site of NDO may play a key role in substrate specificity toward flavanone and isoflavanone. Site-directed mutants of NDO with substitution of Phe224 with Tyr biotransformed only the (S)-stereoisomers of flavanone and isoflavanone, producing an 8-OH group on the A-ring. In contrast, the Phe224Cys and Phe224Gln substitutions, which used (2S)-flavanone as a substrate, and Phe224Lys, which transformed (2S)-flavanone and (3S)-isoflavanone, each showed lower activity than the Phe224Tyr substitution. The remainder of the tested mutants had no activity with flavanone and isoflavanone. Protein docking studies of flavanone and isoflavanone to the modeled mutant enzyme structures revealed that an expanded substrate binding site, due to mutation at 224, as well as appropriate hydrophobic interaction with the residue at 224, are critical for successful binding of the substrates. Results of this study also suggested that in addition to the previously known Phe352, the Phe224 site of NDO appears to be important site for expanding the substrate range of NDO and bringing regiospecific and stereospecific hydroxylation reactions to C8 of the flavanone and isoflavanone A-rings.

Authors+Show Affiliations

Department of Environmental Science and Engineering, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

22391970

Citation

Seo, Jiyoung, et al. "Amino Acid Substitutions in Naphthalene Dioxygenase From Pseudomonas Sp. Strain NCIB 9816-4 Result in Regio- and Stereo-specific Hydroxylation of Flavanone and Isoflavanone." Applied Microbiology and Biotechnology, vol. 97, no. 2, 2013, pp. 693-704.
Seo J, Ryu JY, Han J, et al. Amino acid substitutions in naphthalene dioxygenase from Pseudomonas sp. strain NCIB 9816-4 result in regio- and stereo-specific hydroxylation of flavanone and isoflavanone. Appl Microbiol Biotechnol. 2013;97(2):693-704.
Seo, J., Ryu, J. Y., Han, J., Ahn, J. H., Sadowsky, M. J., Hur, H. G., & Chong, Y. (2013). Amino acid substitutions in naphthalene dioxygenase from Pseudomonas sp. strain NCIB 9816-4 result in regio- and stereo-specific hydroxylation of flavanone and isoflavanone. Applied Microbiology and Biotechnology, 97(2), 693-704. https://doi.org/10.1007/s00253-012-3962-y
Seo J, et al. Amino Acid Substitutions in Naphthalene Dioxygenase From Pseudomonas Sp. Strain NCIB 9816-4 Result in Regio- and Stereo-specific Hydroxylation of Flavanone and Isoflavanone. Appl Microbiol Biotechnol. 2013;97(2):693-704. PubMed PMID: 22391970.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Amino acid substitutions in naphthalene dioxygenase from Pseudomonas sp. strain NCIB 9816-4 result in regio- and stereo-specific hydroxylation of flavanone and isoflavanone. AU - Seo,Jiyoung, AU - Ryu,Ji-Young, AU - Han,Jaehong, AU - Ahn,Joong-Hoon, AU - Sadowsky,Michael J, AU - Hur,Hor-Gil, AU - Chong,Youhoon, Y1 - 2012/03/06/ PY - 2012/01/09/received PY - 2012/02/09/accepted PY - 2012/02/08/revised PY - 2012/3/7/entrez PY - 2012/3/7/pubmed PY - 2013/6/25/medline SP - 693 EP - 704 JF - Applied microbiology and biotechnology JO - Appl. Microbiol. Biotechnol. VL - 97 IS - 2 N2 - Wild-type naphthalene dioxygenase (NDO) from Pseudomonas sp. strain NCIB 9816-4 transforms relatively planar flavone and isoflavone to cis-dihydrodiols. However, this enzyme cannot catalyze the transformation of flavanone and isoflavanone in which a phenyl group bonds to the stereogenic C2 or C3 of the C-ring. Protein modeling suggested that Phe224 in the substrate binding site of NDO may play a key role in substrate specificity toward flavanone and isoflavanone. Site-directed mutants of NDO with substitution of Phe224 with Tyr biotransformed only the (S)-stereoisomers of flavanone and isoflavanone, producing an 8-OH group on the A-ring. In contrast, the Phe224Cys and Phe224Gln substitutions, which used (2S)-flavanone as a substrate, and Phe224Lys, which transformed (2S)-flavanone and (3S)-isoflavanone, each showed lower activity than the Phe224Tyr substitution. The remainder of the tested mutants had no activity with flavanone and isoflavanone. Protein docking studies of flavanone and isoflavanone to the modeled mutant enzyme structures revealed that an expanded substrate binding site, due to mutation at 224, as well as appropriate hydrophobic interaction with the residue at 224, are critical for successful binding of the substrates. Results of this study also suggested that in addition to the previously known Phe352, the Phe224 site of NDO appears to be important site for expanding the substrate range of NDO and bringing regiospecific and stereospecific hydroxylation reactions to C8 of the flavanone and isoflavanone A-rings. SN - 1432-0614 UR - https://www.unboundmedicine.com/medline/citation/22391970/Amino_acid_substitutions_in_naphthalene_dioxygenase_from_Pseudomonas_sp__strain_NCIB_9816_4_result_in_regio__and_stereo_specific_hydroxylation_of_flavanone_and_isoflavanone_ L2 - https://dx.doi.org/10.1007/s00253-012-3962-y DB - PRIME DP - Unbound Medicine ER -