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Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.

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Title
Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.
Other Titles
북극 클라미도모나스 유래 오메가-6 지방산 합성효소의 활성 분석 및 특이적 돌연변이를 통한 효소활성 변화 연구
Authors
Jung, Woongsic
Kim, Sanghee
Choi, Han-Gu
Kang, Sung-Ho
Han, Se Jong
Kim, Eun Jae
Subject
Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology
Keywords
Chlamydomonas sp.; Arctic green microalga; fatty acid methyl ester; linoleic acid methyl ester; oleic acid methyl ester; psychrophilic
Issue Date
2017
Citation
Jung, Woongsic, et al. 2017. "Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.". PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY, 47: 143-150.
Abstract
Arctic Chlamydomonas sp. is a dominant microalgal strain in cold or frozen freshwater in the Arctic region. The full-length open reading frame of the omega-6 fatty acid desaturase gene (AChFAD) was obtained from the transcriptomic database of Arctic Chlamydomonas sp. from the KOPRI Culture Collection of Polar Microorganisms (KCCPM). Amino acid sequence analysis indicated the presence of three conserved histidine-rich segments as unique characteristics of omega-6 FADs, and three transmembrane regions transported to plastidic membranes by chloroplast transit peptides in the N-terminal region. The AChFAD6 desaturase activity was examined by expressing wild-type and V254A mutant (Mut-AChFAD6) heterologous recombinant proteins. Quantitative gas chromatography indicated that the concentration of linoleic acids in AChFAD6-transformed cells increased more than three-fold (6.73 ± 0.13 mg g-1 dry cell weight (DCW)) compared with cells transformed with vector alone. In contrast, transformation with Mut-AChFAD6 increased the concentration of oleic acid to 9.23 ± 0.18 mg g-1 DCW, indicating a change in enzymatic activity to mimic that of stearoyl-CoA desaturase (SCD). These results demonstrate that AChFAD6 of Arctic Chlamydomonas sp. increases membrane fluidity by enhancing desaturating C18 fatty acids and facilitates production of large quantities of linoleic fatty acids in prokaryotic expression systems.
URI
http://repository.kopri.re.kr/handle/201206/5811
DOI
http://dx.doi.org/10.1080/10826068.2016.1188311
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