Document Detail


Conjugated fatty acid synthesis: residues 111 and 115 influence product partitioning of Momordica charantia conjugase.
MedLine Citation:
PMID:  22451660     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Conjugated linolenic acids (CLNs), 18:3 Δ(9,11,13), lack the methylene groups found between the double bonds of linolenic acid (18:3 Δ(9,12,15)). CLNs are produced by conjugase enzymes that are homologs of the oleate desaturases FAD2. The goal of this study was to map the domain(s) within the Momordica charantia conjugase (FADX) responsible for CLN formation. To achieve this, a series of Momordica FADX-Arabidopsis FAD2 chimeras were expressed in the Arabidopsis fad3fae1 mutant, and the transformed seeds were analyzed for the accumulation of CLN. These experiments identified helix 2 and the first histidine box as a determinant of conjugase product partitioning into punicic acid (18:3 Δ(9cis,11trans,13cis)) or α-eleostearic acid (18:3 Δ(9cis,11trans,13trans)). This was confirmed by analysis of a FADX mutant containing six substitutions in which the sequence of helix 2 and first histidine box was converted to that of FAD2. Each of the six FAD2 substitutions was individually converted back to the FADX equivalent identifying residues 111 and 115, adjacent to the first histidine box, as key determinants of conjugase product partitioning. Additionally, expression of FADX G111V and FADX G111V/D115E resulted in an approximate doubling of eleostearic acid accumulation to 20.4% and 21.2%, respectively, compared with 9.9% upon expression of the native Momordica FADX. Like the Momordica conjugase, FADX G111V and FADX D115E produced predominantly α-eleostearic acid and little punicic acid, but the FADX G111V/D115E double mutant produced approximately equal amounts of α-eleostearic acid and its isomer, punicic acid, implicating an interactive effect of residues 111 and 115 in punicic acid formation.
Authors:
Richa Rawat; Xiao-Hong Yu; Marie Sweet; John Shanklin
Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, Non-P.H.S.     Date:  2012-03-26
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  287     ISSN:  1083-351X     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2012 May 
Date Detail:
Created Date:  2012-05-14     Completed Date:  2012-07-13     Revised Date:  2013-06-26    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  16230-7     Citation Subset:  IM    
Affiliation:
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, New York 11794, USA.
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MeSH Terms
Descriptor/Qualifier:
Arabidopsis / enzymology,  genetics
Linolenic Acids / biosynthesis*,  genetics
Momordica charantia / enzymology*,  genetics
Plant Proteins / genetics,  metabolism*
Plants, Genetically Modified / enzymology,  genetics
gamma-Glutamyl Hydrolase / genetics,  metabolism*
Chemical
Reg. No./Substance:
0/Linolenic Acids; 0/Plant Proteins; 0/punicic acid; EC 3.4.19.9/gamma-Glutamyl Hydrolase
Comments/Corrections

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