中国精品科技期刊
中国高校百佳科技期刊
中国中文核心期刊
中国科学引文数据库核心期刊

PDF
Cite
Share

facebook

twitter

google

linkedin

Journal of China Pharmaceutical University > 2021 > 52(6): 742-750. > DOI: 10.11665/j.issn.1000-5048.20210613

ZHANG Zhan, FENG Yan, LI Qian, CUI Li. Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii[J]. Journal of China Pharmaceutical University, 2021, 52(6): 742-750. DOI: 10.11665/j.issn.1000-5048.20210613

Citation:

ZHANG Zhan, FENG Yan, LI Qian, CUI Li. Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii[J]. Journal of China Pharmaceutical University, 2021, 52(6): 742-750. DOI: 10.11665/j.issn.1000-5048.20210613

ZHANG Zhan, FENG Yan, LI Qian, CUI Li. Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii[J]. Journal of China Pharmaceutical University, 2021, 52(6): 742-750. DOI: 10.11665/j.issn.1000-5048.20210613

Citation:

ZHANG Zhan, FENG Yan, LI Qian, CUI Li. Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii[J]. Journal of China Pharmaceutical University, 2021, 52(6): 742-750. DOI: 10.11665/j.issn.1000-5048.20210613

PDF (2813 KB)

Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii

1.
State Key Laboratory of Microbial Metabolism, School of Life Science and Technology, Shanghai Jiao Tong University, Shanghai 200240
2.
School of Life Science & Technology, China Pharmaceutical University, Nanjing 210009, China

Funds: This study was supported by the National Natural Science Foundation of China (No.21977067, No.31770098, No.31620103901) and the National Key R&D Program of China (No.2018YFA0900702)

More Information

Received Date: April 21, 2021
Revised Date: November 03, 2021

Graphical Abstract

Abstract

The purpose of this article is to express the UDP-glucose 4-epimerase from Acinetobacter baumannii AB0057, characterize its enzymatic properties and analyze its structure and function.The epimerase gene was constructed into pET-28a expression vector and heterologously expressed in BL21(DE3).Enzyme activity was assayed using high performance liquid chromatography.The structure and key residues were analyzed by phylogenetic analysis, sequence alignment, homology modeling and molecular docking.Results indicated that the recombinant enzyme Gne1 was expressed at a molecular weight of 38.9 kD, with an optimum temperature of 44 °C and an optimum pH of 6.0 .Michaelis-Menten parameters K_M and k_cat were (1.227 ± 0.082 4) mmol/L and (82.64 ± 3.562) × 10^-3?min^-1, respectively.This enzyme belongs to NADB_Rossmann superfamily and UDP_G4E_1_SDR_e subgroup with typical GXGXXG and YXXXK sequence motifs.The N-terminal structural domain bound to NAD, while the C-terminal structural domain bound to substrate, and the catalytic key sites were S125 and Y150.The current work verified the epimerase activity of Gne1, explained its sequence and structural features, revealed its binding mode with substrates and cofactors, and analyzed the key residues, which provides a basis for protein engineering to improve the epimerase activity and then use biological enzymatic method to synthesize rare functional sugars.
- epimerase,
- enzymatic property,
- sequence and structure analysis,
- synthesis of rare functional sugar

FullText(HTML)

References (21)

References

[1]	. J Biosci Bioeng，2004，97（2）：89-94.
[2]	Mu WM，Zhang WL，Feng YH，et al. Recent advances on applications and biotechnological production of D-psicose［J］. Appl Microbiol Biotechnol，2012，94（6）：1461-1467.
[3]	Kim P. Current studies on biological tagatose production using L-arabinose isomerase：a review and future perspective［J］. Appl Microbiol Biotechnol，2004，65（3）：243-249.
[4]	Oh DK. Tagatose：properties，applications，and biotechnological processes［J］. Appl Microbiol Biotechnol，2007，76（1）：1-8.
[5]	Jenkinson SF，Fleet GW，Nash RJ，et al. Looking-glass synergistic pharmacological chaperones：DGJ and L-DGJ from the enantiomers of tagatose［J］. Org Lett，2011，13（15）：4064-4067.
[6]	Izumori K. Izumoring：a strategy for bioproduction of all hexoses［J］. J Biotechnol，2006，124（4）：717-722.
[7]	Zhu HM，Sun B，Li YJ，et al. KfoA，the UDP-glucose-4-epimerase of Escherichia coli strain O5：K4：H4，shows preference for acetylated substrates［J］. Appl Microbiol Biotechnol，2018，102（2）：751-761.
[8]	Ishiyama N，Creuzenet C，Lam JS，et al. Crystal structure of WbpP，a genuine UDP-N-acetylglucosamine 4-epimerase from Pseudomonas aeruginosa：substrate specificity in udp-hexose 4-epimerases［J］. J Biol Chem，2004，279（21）：22635-22642.
[9]	Frey PA. The Leloir pathway：a mechanistic imperative for three enzymes to change the stereochemical configuration of a single carbon in galactose［J］. FASEB J，1996，10（4）：461-470.
[10]	Holden HM，Rayment I，Thoden JB. Structure and function of enzymes of the Leloir pathway for galactose metabolism［J］. J Biol Chem，2003，278（45）：43885-43888.
[11]	Ostash B，Doud EH，Lin C，et al. Complete characterization of the seventeen step moenomycin biosynthetic pathway［J］. Biochemistry，2009，48（37）：8830-8841.
[12]	Song HB，He M，Cai ZP，et al. UDP-glucose 4-epimerase and β-1，4-galactosyltransferase from the oyster Magallana gigas as valuable biocatalysts for the production of galactosylated products［J］. Int J Mol Sci，2018，19（6）：1600-1610.
[13]	Kim HJ，Kang SY，Park JJ，et al. Novel activity of UDP-galactose-4-epimerase for free monosaccharide and activity improvement by active site-saturation mutagenesis［J］. Appl Biochem Biotechnol，2011，163（3）：444-451.
[14]	Wilson DB，Hogness DS. The enzymes of the galactose operon in Escherichia coli. I. purification and characterization of uridine diphosphogalactose 4-epimerase［J］. J Biol Chem，1964，239：2469-2481.
[15]	Swanson BA，Frey PA. Identification of lysine 153 as a functionally important residue in UDP-galactose 4-epimerase from Escherichia coli［J］. Biochemistry，1993，32（48）：13231-13236.
[16]	Liu Y，Thoden JB，Kim J，et al. Mechanistic roles of tyrosine 149 and serine 124 in UDP-galactose 4-epimerase from Escherichia coli［J］. Biochemistry，1997，36（35）：10675-10684.
[17]	Liu Y，Vanhooke JL，Frey PA. UDP-galactose 4-epimerase：NAD⁺ content and a charge-transfer band associated with the substrate-induced conformational transition［J］. Biochemistry，1996，35（23）：7615-7620.
[18]	Frey PA，Hegeman AD. Chemical and stereochemical actions of UDP-galactose 4-epimerase［J］. Acc Chem Res，2013，46（7）：1417-1426.
[19]	Thoden JB，Hegeman AD，Wesenberg G，et al. Structural analysis of UDP-sugar binding to UDP-galactose 4-epimerase from Escherichia coli［J］. Biochemistry，1997，36（21）：6294-6304.
[20]	Thoden JB，Holden HM. Dramatic differences in the binding of UDP-galactose and UDP-glucose to UDP-galactose 4-epimerase from Escherichia coli［J］. Biochemistry，1998，37（33）：11469-11477.
[21]	Thoden JB，Frey PA，Holden HM. Molecular structure of the NADH/UDP-glucose abortive complex of UDP-galactose 4-epimerase from Escherichia coli：implications for the catalytic mechanism［J］. Biochemistry，1996，35（16）：5137-5144.

Related Articles

Related Articles

[1]	TAO Junjie, YANG Jie, WEN Xiaodong. Studies on the chemical constituents in the aerial parts of Paris polyphylla var. Yunnanensis[J]. Journal of China Pharmaceutical University, 2020, 51(4): 454-460. DOI: 10.11665/j.issn.1000-5048.20200410
[2]	WANG Xiaoying, WANG Xiaying, QIU Liangzhen, OUYANG Huizhi, XU Wei. Cytotoxicity and cellular uptake of paclitaxel-loaded carboxymethyl chitosan-rhein polymeric micelles in MCF-7 cells[J]. Journal of China Pharmaceutical University, 2020, 51(1): 33-37. DOI: 10.11665/j.issn.1000-5048.20200106
[3]	CHENG Weihua, WANG Wenqian, SHANG Hai, ZHANG Hongwu, GUO Qiang, CHEN Hong, ZOU Zhongmei. Synthesis and cytotoxicity study of cembrane triazole derivatives[J]. Journal of China Pharmaceutical University, 2018, 49(1): 56-63. DOI: 10.11665/j.issn.1000-5048.20180108
[4]	TANG Yong, LIAO Shengrong, LI Jinsheng, LIU Yonghong. Design, synthesis and cytotoxic activities of fluorine-containing 2, 5-diketopiperazine derivatives[J]. Journal of China Pharmaceutical University, 2016, 47(4): 412-421. DOI: 10.11665/j.issn.1000-5048.20160405
[5]	SUN Hao, YANG Jiapei, MAO Yong, WANG Dandan, YU Feng. Involvement of Fas-dependent pathway in rhein-induced apoptosis of HK-2 cells[J]. Journal of China Pharmaceutical University, 2015, 46(4): 469-475. DOI: 10.11665/j.issn.1000-5048.20150414
[6]	WU Mingming, FANG Lei, GOU Shaohua, CHEN Li. 以2-甲基-2-取代苯氧基丙酸为离去基团的铂(Ⅱ)配合物的合成、表征及细胞毒活性[J]. Journal of China Pharmaceutical University, 2013, 44(4): 303-306. DOI: 10.11665/j.issn.1000-5048.20130403
[7]	FEI Jia, SUN Min-jie, PING Qi-neng, ZHANG Can. Preparation,characterization and safety evaluation for N-octyl-N-quaternary ammonium chitosan/CMCNa composite micelles[J]. Journal of China Pharmaceutical University, 2012, 43(4): 318-324.
[8]	SHEN Yan, LI Qian, FAN Da, LI Rui-jun, TU Jia-sheng. Influencing factors of hydrophobic modified hyaluronic acid-based cationic micelles and cell targeting in siRNA transfection[J]. Journal of China Pharmaceutical University, 2012, 43(2): 124-129.
[9]	GUO Zhong, ZHAO Jin, MA Jian-xiu, FU Si-wu. Effects of toxin A from Clostridium difficile on SMMC-7721 cells proliferation and apoptosis[J]. Journal of China Pharmaceutical University, 2009, 40(3): 250-253.
[10]	Structure-Cytotoxic Activity Relationships of Diterpenoid Tanshinones from Salvias[J]. Journal of China Pharmaceutical University, 2002, (1): 8-14.

Get Citation

PDF

XML

Article views (117) PDF downloads (335)

Turn off MathJax

Article Contents

Abstract

References

Related Articles

/

DownLoad: Full-Size Img PowerPoint

Return