Evolution-related amino acids play important role in determining regioselectivity of fatty acid desaturase from <Emphasis Type="Italic">Pichia pastoris</Emphasis> |
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Authors: | Xinxin Zhang Dongsheng Wei Mingchun Li Yuyan Qi Laijun Xing |
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Institution: | (1) Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Department of Microbiology, Nankai University, Tianjin, 300071, P.R. China |
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Abstract: | ω3-fatty acid desaturase and Δ12-fatty acid desaturase of Pichia pastoris with distinguishable regioselectivity and high degree of sequence similarity were chosen for regioselectivity research. Chimeras
were constructed in which Histidine-rich boxes 1, 2 and the carboxyl terminal region of ω3-fatty acid desaturase were replaced with corresponding region of Δ12-fatty acid desaturase. The replacement was found to result in a change of regioselectivity from ωy to x + 3 by functionally characterizing these chimeric enzymes in Saccharomyces cerevisae strain INVScI. Using site-directed mutagenesis, we further demonstrated that seven conserved amino acids of ω3-fatty acid desaturase within the first two Histidine-rich regions are responsible for the regioselectivity switch. Therefore,
the regioselectivity of fatty acid desaturases may be better understood by investigating the evolutionary relationships of
different fatty acid desaturases.
Dongsheng Wei is the partake of first-author’s profits. |
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Keywords: | Pichia pastoris ω 3-Fatty acid desaturase Δ 12-Fatty acid desaturase Chimeras Site-directed mutagenesis Regioselectively |
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