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Title: Castor Stearoyl-ACP Desaturase Can Synthesize a Vicinal Diol by Dioxygenase Chemistry

Abstract

In previous work, we identified a triple mutant of the castor (Ricinus communis) stearoyl-Acyl Carrier Protein desaturase (T117R/G188L/D280K) that, in addition to introducing a double bond into stearate to produce oleate, performed an additional round of oxidation to convert oleate to a trans allylic alcohol acid. To determine the contributions of each mutation, in the present work we generated individual castor desaturase mutants carrying residue changes corresponding to those in the triple mutant and investigated their catalytic activities. We observed that T117R, and to a lesser extent D280K, accumulated a novel product, namely erythro-9, 10-dihydroxystearate, that we identified via its methyl ester through gas chromatography/mass spectrometry and comparison with authentic standards. The use of 18O2 labeling showed that the oxygens of both hydroxyl moieties originate from molecular oxygen rather than water. Incubation with an equimolar mixture of 18O2 and 18O2 demonstrated that both hydroxyl oxygens originate from a single molecule of O2, proving the product is the result of dioxygenase catalysis. Using prolonged incubation, we discovered that wild-type castor desaturase is also capable of forming erythro-9, 10-dihydroxystearate, which presents a likely explanation for its accumulation to approximately 0.7% in castor oil, of which the biosynthetic origin had remained enigmatic formore » decades. In summary, the findings presented here expand the documented constellation of diiron enzyme catalysis to include a dioxygenase reactivity in which an unactivated alkene is converted to a vicinal diol.« less

Authors:
ORCiD logo [1]; ORCiD logo [1]; ORCiD logo [1]; ORCiD logo [1]; ORCiD logo [2]; ORCiD logo [1]
  1. Biology Department, Brookhaven National Laboratory, Upton, New York 11973
  2. Department of Chemistry, Carleton University, Ottawa, Ontario, Canada K1S 5B6
Publication Date:
Research Org.:
Brookhaven National Lab. (BNL), Upton, NY (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES)
OSTI Identifier:
1577374
Alternate Identifier(s):
OSTI ID: 1579893
Report Number(s):
BNL-212399-2019-JAAM
Journal ID: ISSN 0032-0889; /plantphysiol/182/2/730.atom
Grant/Contract Number:  
SC0012704
Resource Type:
Published Article
Journal Name:
Plant Physiology (Bethesda)
Additional Journal Information:
Journal Name: Plant Physiology (Bethesda) Journal Volume: 182 Journal Issue: 2; Journal ID: ISSN 0032-0889
Publisher:
Oxford University Press
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Whittle, Edward J., Cai, Yuanheng, Keereetaweep, Jantana, Chai, Jin, Buist, Peter H., and Shanklin, John. Castor Stearoyl-ACP Desaturase Can Synthesize a Vicinal Diol by Dioxygenase Chemistry. United States: N. p., 2019. Web. doi:10.1104/pp.19.01111.
Whittle, Edward J., Cai, Yuanheng, Keereetaweep, Jantana, Chai, Jin, Buist, Peter H., & Shanklin, John. Castor Stearoyl-ACP Desaturase Can Synthesize a Vicinal Diol by Dioxygenase Chemistry. United States. doi:10.1104/pp.19.01111.
Whittle, Edward J., Cai, Yuanheng, Keereetaweep, Jantana, Chai, Jin, Buist, Peter H., and Shanklin, John. Thu . "Castor Stearoyl-ACP Desaturase Can Synthesize a Vicinal Diol by Dioxygenase Chemistry". United States. doi:10.1104/pp.19.01111.
@article{osti_1577374,
title = {Castor Stearoyl-ACP Desaturase Can Synthesize a Vicinal Diol by Dioxygenase Chemistry},
author = {Whittle, Edward J. and Cai, Yuanheng and Keereetaweep, Jantana and Chai, Jin and Buist, Peter H. and Shanklin, John},
abstractNote = {In previous work, we identified a triple mutant of the castor (Ricinus communis) stearoyl-Acyl Carrier Protein desaturase (T117R/G188L/D280K) that, in addition to introducing a double bond into stearate to produce oleate, performed an additional round of oxidation to convert oleate to a trans allylic alcohol acid. To determine the contributions of each mutation, in the present work we generated individual castor desaturase mutants carrying residue changes corresponding to those in the triple mutant and investigated their catalytic activities. We observed that T117R, and to a lesser extent D280K, accumulated a novel product, namely erythro-9, 10-dihydroxystearate, that we identified via its methyl ester through gas chromatography/mass spectrometry and comparison with authentic standards. The use of 18O2 labeling showed that the oxygens of both hydroxyl moieties originate from molecular oxygen rather than water. Incubation with an equimolar mixture of 18O2 and 18O2 demonstrated that both hydroxyl oxygens originate from a single molecule of O2, proving the product is the result of dioxygenase catalysis. Using prolonged incubation, we discovered that wild-type castor desaturase is also capable of forming erythro-9, 10-dihydroxystearate, which presents a likely explanation for its accumulation to approximately 0.7% in castor oil, of which the biosynthetic origin had remained enigmatic for decades. In summary, the findings presented here expand the documented constellation of diiron enzyme catalysis to include a dioxygenase reactivity in which an unactivated alkene is converted to a vicinal diol.},
doi = {10.1104/pp.19.01111},
journal = {Plant Physiology (Bethesda)},
number = 2,
volume = 182,
place = {United States},
year = {2019},
month = {12}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
DOI: 10.1104/pp.19.01111

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Cited by: 1 work
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