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A Simple Enzymatic Method for Quantitation of 2’-Fucosyllactose
( Emine Seydametova ),( Jonghyeok Shin ),( Jiwon Yu ),( Dae-hyuk Kweon ) 한국미생물생명공학회(구 한국산업미생물학회) 2018 Journal of microbiology and biotechnology Vol.28 No.7
2’-Fucosyllactose (2’-FL) is one of the most important human milk oligosaccharides and has several health benefits for infants. The levels of 2’-FL in breast milk or samples from other sources can be quantified by high-performance liquid chromatography. However, this method cannot be used for simultaneous detection of the target compound in numerous samples. Here, we developed a simple method for quantifying 2’-FL in a microplate format. The method involves two steps: (i) release of L-fucose from 2’-FL by α-(1-2,3,4,6)-L-fucosidase and (ii) measurement of NADPH formed during the oxidation of L-fucose by L-fucose dehydrogenase. This method enables measurement of up to 5 g/l 2’-FL in 50 min using a 96- well microplate. The efficiency and simplicity of the proposed method make it suitable for the analyses of a large number of samples simultaneously.
Yu, Jiwon,Shin, Jonghyeok,Park, Myungseo,Seydametova, Emine,Jung, Sang-Min,Seo, Jin-Ho,Kweon, Dae-Hyuk Elsevier 2018 Metabolic engineering Vol.48 No.-
<P><B>Abstract</B></P> <P>Fucosyllactoses (FLs), present in human breast milk, have been reported to benefit human health immensely. Especially, 3-fucosyllactose (3-FL) has numerous benefits associated with a healthy gut ecosystem. Metabolic engineering of microorganisms is thought to be currently the only option to provide an economically feasible route for large-scale production of 3-FL. However, engineering principles for α-1,3-fucosyltransferases (1,3-FTs) are not well-known, resulting in the lower productivity of 3-FL than that of 2′-fucosyllactose (2′-FL), although both 2′-FL and 3-FL follow a common pathway to produce GDP-<SMALL>L</SMALL>-fucose. The C-terminus of 1,3-FTs is composed of heptad repeats, responsible for dimerization of the enzymes, and a peripheral membrane anchoring region. It has long been thought that truncation of most heptad repeats, retaining just 1 or 2, helps the soluble expression of 1,3-FTs. However, whether the introduction of truncated version of 1,3-FTs enhances the production of 3-FL in a metabolically engineered strain, is yet to be tested. In this study, the effect of these structural components on the production of 3-FL in <I>Escherichia coli</I> was evaluated through systematic truncation and elongation of the C-terminal regions of three 1,3-FTs from <I>Helicobacter pylori</I>. Although these three 1,3-FTs contained heptad repeats and membrane-anchoring regions of varying lengths, they commonly exhibited an optimal performance when the number of heptad repeats was increased, and membrane-binding region was removed. The production of 3-FL could be increased 10–20-fold through this simple strategy.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Heptad repeats in several α-1,3-fucosyltransferases were truncated or elongated. </LI> <LI> Dimerization of 1,3-FT was essential for enzyme activity and thermostability. </LI> <LI> 3-fucosyllactose was better produced in <I>E. coli</I> with longer heptad repeats. </LI> </UL> </P>
Stabilization of Proteins Inside Cells by Immobilization on Plasmid
Yunjeong PARK,Jonghyeok SHIN,Chakhee KIM,Hooyeon KIM,Emine SEYDAMETOVA,Dae-Hyuk KWEON,Choongjin BAN,Joon-Bum PARK,Myungseo PARK,Younghun JUNG,Seokoh MOON,Seok-Hyeon YU,Sora CHO,Hye Rin KIM,Hyun Seok O 한국생물공학회 2018 한국생물공학회 학술대회 Vol.2018 No.10
Transdermal delivery of myricetin for the hyperhidrosis treatment using lipid nanoparticles
Choongjin BAN,Joon-Bum PARK,Sora CHO,Hyerin KIM,Jonghyeok SHIN,Chakhee KIM,Hooyeon KIM,Yunjeong PARK,Myungseo PARK,Younghun JUNG,Seokoh MOON,Emine SEYDAMETOVA,Seok-Hyeon YU,Dae-Hyuk KWEON 한국생물공학회 2018 한국생물공학회 학술대회 Vol.2018 No.10