Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus

Research output: Contribution to journal › Journal article › Research › peer-review

Standard

Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus. / Zhang, Yan; Eser, Bekir Engin; Kougioumtzoglou, Georgios; Eser, Zekiye; Poborsky, Michal; Kishino, Shigenobu; Takeuchi, Michiki; Ogawa, Jun; Kristensen, Peter; Guo, Zheng.

In: Biochemical Engineering Journal, Vol. 171, 108006, 2021.

Research output: Contribution to journal › Journal article › Research › peer-review

Harvard

Zhang, Y, Eser, BE, Kougioumtzoglou, G, Eser, Z, Poborsky, M, Kishino, S, Takeuchi, M, Ogawa, J, Kristensen, P & Guo, Z 2021, 'Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus', Biochemical Engineering Journal, vol. 171, 108006. https://doi.org/10.1016/j.bej.2021.108006

APA

Zhang, Y., Eser, B. E., Kougioumtzoglou, G., Eser, Z., Poborsky, M., Kishino, S., Takeuchi, M., Ogawa, J., Kristensen, P., & Guo, Z. (2021). Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus. Biochemical Engineering Journal, 171, [108006]. https://doi.org/10.1016/j.bej.2021.108006

Vancouver

Zhang Y, Eser BE, Kougioumtzoglou G, Eser Z, Poborsky M, Kishino S et al. Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus. Biochemical Engineering Journal. 2021;171. 108006. https://doi.org/10.1016/j.bej.2021.108006

Author

Zhang, Yan ; Eser, Bekir Engin ; Kougioumtzoglou, Georgios ; Eser, Zekiye ; Poborsky, Michal ; Kishino, Shigenobu ; Takeuchi, Michiki ; Ogawa, Jun ; Kristensen, Peter ; Guo, Zheng. / Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus. In: Biochemical Engineering Journal. 2021 ; Vol. 171.

Bibtex

@article{d23dc9ffb9da424398c889427b8dccc9,
title = "Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus",
abstract = "Fatty acid hydratase (FAH) mediated hydroxy fatty acid (HFA) production is a promising enzymatic route that demands diversification of hydration position to access a broader range of high-value HFAs. FA-HY1 is a promiscuous FAH from Lactobacillus Acidophilus, whereas its homolog from the same organism, FA-HY2, is strict in substrate scope and regioselectivity. Our earlier work demonstrated that three amino acid mutations at the carboxylate end of the substrate (T391/H393/I378 in FA-HY2) shift regioselectivity of FA-HY2 towards that of FA-HY1. Here, we explore alanine 216 of FA-HY2 as a hot-spot residue at the omega end of the substrate. A quadruple mutant (T391S/H393S/I378 P/A216S) demonstrates further shift in regioselectivity towards FA-HY1. Moreover, site-saturation mutagenesis of this residue in FA-HY1 (S218) led to novel variants exhibiting significant changes in regioselectivity for EPA (eicosapentaenoic acid) as substrate, where, unlike wild-type enzyme, 15-OH product was the dominant product (63:37 for wild-type vs. 26:74 for S218I mutant; 12-OH:15-OH). Alterations in conversion levels that indicate pronounced correlation to the exchanged residue type were also detected. A likely explanation for the observed differences is provided based on structural, statistical and kinetic analysis.",
keywords = "Enzyme engineering, Fatty acid hydratase, Hydroxy fatty acid, Regioselectivity, Site saturation mutagenesis",
author = "Yan Zhang and Eser, {Bekir Engin} and Georgios Kougioumtzoglou and Zekiye Eser and Michal Poborsky and Shigenobu Kishino and Michiki Takeuchi and Jun Ogawa and Peter Kristensen and Zheng Guo",
note = "Publisher Copyright: {\textcopyright} 2021 Elsevier B.V.",
year = "2021",
doi = "10.1016/j.bej.2021.108006",
language = "English",
volume = "171",
journal = "Biochemical Engineering Journal",
issn = "1369-703X",
publisher = "Elsevier",

}

RIS

TY - JOUR

T1 - Effects of the engineering of a single binding pocket residue on specificity and regioselectivity of hydratases from Lactobacillus Acidophilus

AU - Zhang, Yan

AU - Eser, Bekir Engin

AU - Kougioumtzoglou, Georgios

AU - Eser, Zekiye

AU - Poborsky, Michal

AU - Kishino, Shigenobu

AU - Takeuchi, Michiki

AU - Ogawa, Jun

AU - Kristensen, Peter

AU - Guo, Zheng

N1 - Publisher Copyright: © 2021 Elsevier B.V.

PY - 2021

Y1 - 2021

N2 - Fatty acid hydratase (FAH) mediated hydroxy fatty acid (HFA) production is a promising enzymatic route that demands diversification of hydration position to access a broader range of high-value HFAs. FA-HY1 is a promiscuous FAH from Lactobacillus Acidophilus, whereas its homolog from the same organism, FA-HY2, is strict in substrate scope and regioselectivity. Our earlier work demonstrated that three amino acid mutations at the carboxylate end of the substrate (T391/H393/I378 in FA-HY2) shift regioselectivity of FA-HY2 towards that of FA-HY1. Here, we explore alanine 216 of FA-HY2 as a hot-spot residue at the omega end of the substrate. A quadruple mutant (T391S/H393S/I378 P/A216S) demonstrates further shift in regioselectivity towards FA-HY1. Moreover, site-saturation mutagenesis of this residue in FA-HY1 (S218) led to novel variants exhibiting significant changes in regioselectivity for EPA (eicosapentaenoic acid) as substrate, where, unlike wild-type enzyme, 15-OH product was the dominant product (63:37 for wild-type vs. 26:74 for S218I mutant; 12-OH:15-OH). Alterations in conversion levels that indicate pronounced correlation to the exchanged residue type were also detected. A likely explanation for the observed differences is provided based on structural, statistical and kinetic analysis.

AB - Fatty acid hydratase (FAH) mediated hydroxy fatty acid (HFA) production is a promising enzymatic route that demands diversification of hydration position to access a broader range of high-value HFAs. FA-HY1 is a promiscuous FAH from Lactobacillus Acidophilus, whereas its homolog from the same organism, FA-HY2, is strict in substrate scope and regioselectivity. Our earlier work demonstrated that three amino acid mutations at the carboxylate end of the substrate (T391/H393/I378 in FA-HY2) shift regioselectivity of FA-HY2 towards that of FA-HY1. Here, we explore alanine 216 of FA-HY2 as a hot-spot residue at the omega end of the substrate. A quadruple mutant (T391S/H393S/I378 P/A216S) demonstrates further shift in regioselectivity towards FA-HY1. Moreover, site-saturation mutagenesis of this residue in FA-HY1 (S218) led to novel variants exhibiting significant changes in regioselectivity for EPA (eicosapentaenoic acid) as substrate, where, unlike wild-type enzyme, 15-OH product was the dominant product (63:37 for wild-type vs. 26:74 for S218I mutant; 12-OH:15-OH). Alterations in conversion levels that indicate pronounced correlation to the exchanged residue type were also detected. A likely explanation for the observed differences is provided based on structural, statistical and kinetic analysis.

KW - Enzyme engineering

KW - Fatty acid hydratase

KW - Hydroxy fatty acid

KW - Regioselectivity

KW - Site saturation mutagenesis

U2 - 10.1016/j.bej.2021.108006

DO - 10.1016/j.bej.2021.108006

M3 - Journal article

AN - SCOPUS:85103708005

VL - 171

JO - Biochemical Engineering Journal

JF - Biochemical Engineering Journal

SN - 1369-703X

M1 - 108006

ER -

ID: 272123120