Lipases possess unique genus features during bacterial evolution. Discovery of new thermophilic lipases represents an important field for lipase research and application. The current methods of isolation and identification of new thermophilic lipases from thermophilic microbial fermentation products and through metagenomic methods are laborious. Based on bioinformatics methods and heterologous expression techniques, the Caldisericum exile genomic data from marine hot springs were screened and evolutionarily analyzed, and it was found that BAL81435.1 (GenBank ID) may encode new thermophilic lipase. The sequence was cloned into pET28(a) vector, and the properties of recombinant lipase were characterized after purification. BAL81435.1 encodes a new thermophilic lipase whose substrates are long-chain (>C12) para-nitrophenyl phosphate (p-NPP), with maximum specificity detected at chain length of C16. Its optimal pH was 8.8 and optimal temperature was 60 ℃. This lipase is thermally-stable between 4 ℃ and 60 ℃ and tolerant to most organic solvents, metal ions, denaturing agents and high salinity, but surfactant-sensitive. This study discovers a novel thermophilic lipase encoded by BAL81435.1, which lays foundation for prospective lipase study and exploitation.
LIU Jiejing
,
WANG Minting
,
ZHOU Yuting
,
CHEN Ping
,
OUYANG Yongchang
. Discovery and characterization of a novel thermophilic lipase derived from Caldisericum exile[J]. Food and Fermentation Industries, 2023
, 49(2)
: 41
-46
.
DOI: 10.13995/j.cnki.11-1802/ts.030896
[1] DEMIRJIAN D C, MORIS-VARAS F, CASSIDY C S.Enzymes from extremophiles[J].Current Opinion in Chemical Biology, 2001, 5(2):144-151.
[2] ATALAH J, CACERES-MORENO P, ESPINA G, et al.Thermophiles and the applications of their enzymes as new biocatalysts[J].Bioresource Technology,2019, 280:478-488.
[3] DHEEMAN D S, FRIAS J M, HENEHAN G T.Influence of cultivation conditions on the production of a thermostable extracellular lipase from Amycolatopsis mediterranei DSM 43304[J].Journal of Industrial Microbiology & Biotechnology, 2010, 37(1):1-17.
[4] ZOTTIG X, MEDDEB-MOUELHI F, CHARBONNEAU D M, et al.Characterization of a novel alkalophilic lipase from Aneurinibacillusther moaerophilus:Lid heterogeneity and assignment to family I.5[J].The Protein Journal, 2017, 36(6):478-488.
[5] JO E, KIM J, LEE A, et al.Identification and characterization of a novel thermostable GDSL-Type lipase from Geobacillusther mocatenulatus[J].Journal of Microbiology and Biotechnology, 2021, 31(3):483-491.
[6] ABDEL-FATTAH Y R, GABALLA A A.Identification and over-expression of a thermostable lipase from Geobacillusther moleovorans Toshki in Escherichia coli[J].Microbiological Research,2008, 163(1):13-20.
[7] BANCERZ R, GINALSKA G.A novel thermostable lipase from basidiomycete Bjerkandera adusta R59:Characterisation and esterification studies[J].Journal of Industrial Microbiology & Biotechnology, 2007, 34(8):553-560.
[8] DE FÁTIMA SILVA LOPES M, et al.Characterization of a highly thermostable extracellular lipase from Lactobacillus plantarum[J].International Journal of Food Microbiology, 2002, 76(1/2):107-115.
[9] XIE W, KHOSASIH V, SUWANTO A, et al.Characterization of lipases from Staphylococcus aureus and Staphylococcus epidermidis isolated from human facial sebaceous skin[J].Journal of Microbiology and Biotechnology,2012, 22(1):84-91.
[10] H-KITTIKUN A, PRASERTSAN P, ZIMMERMANN W, et al.Sugar ester synthesis by thermostable lipase from Streptomyces thermocarboxydus ME168[J].Applied Biochemistry and Biotechnology,2012, 166(8):1 969-1 982.
[11] ROYTER M, SCHMIDT M, ELEND C, et al.Thermostable lipases from the extreme thermophilic anaerobic bacteria Thermoanaerobacter thermohydrosulfuricus SOL1 and Caldanaerobacter subterraneus subsp.tengcongensis[J].Extremophiles:Life under Extreme Conditions,2009, 13(5):769-783.
[12] SALAMEH M A, WIEGEL J.Purification and characterization of two highly thermophilic alkaline lipases from Thermosyntropha lipolytica[J].Applied and Environmental Microbiology,2007, 73(23):7 725-7 731.
[13] LÓPEZ-LÓPEZ O, CERDÁNÁM E, GONZÁLEZ SISO M I.New extremophilic lipases and esterases from metagenomics[J].Current Protein & Peptide Science,2014, 15(5):445-455.
[14] TIRAWONGSAROJ P, SRIPRANG R, HARNPICHARNCHAI P, et al.Novel thermophilic and thermostable lipolytic enzymes from a Thailand hot spring metagenomic library[J].Journal of Biotechnology,2008, 133(1):42-49.
[15] 周艳杰,耿鹏,时祎,等.嗜热玫瑰红球菌嗜热脂肪酶的重组表达与酶学性质研究[J].食品与发酵工业,2021,47(13):16-22.
ZHOU Y J, GENG P, SHI Y, et al.Expression and characterization of a novel thermostable lipase from Thermomicrobium roseum[J].Food and Fermentation Industries, 2021,47(13):16-22.
[16] 郭阳阳. 新型嗜热脂解酶的克隆表达及其酶学性质研究[D].无锡:江南大学, 2017.
GUO Y Y.Cloning, expression and characterization of novel thermophilic lipolytic enzymes[D].Wuxi:Jiangnan University, 2017.
[17] ABDELMONAEM M, HATEM B, IMEN G, et al.Classification of EC 3.1.1.3 bacterial true lipases using phylogenetic analysis[J].African Journal of Biotechnology,2010, 9(48):8 243-8 247.
[18] YADAVA K, DUBEYA K, YADAV S, et al.Amino acid sequences based phylogenetic and motif assessment of lipases from different organisms[J].Online Journal of Bioinformatics,2012, 13(3):400-417.
[19] MORI K, YAMAGUCHI K, SAKIYAMA Y, et al.Caldisericum exile gen.nov., sp.nov., an anaerobic, thermophilic, filamentous bacterium of a novel bacterial Phylum, Caldiserica phyl.nov., originally called the candidate phylum OP5, and description of Caldisericaceae fam.nov., Caldisericales ord.nov.and Caldisericia classis nov[J].International Journal of Systematic and Evolutionary Microbiology,2009, 59(11):2 894-2 898.
[20] TAYYAB M, RASHID N, AKHTAR M.Isolation and identification of lipase producing thermophilic Geobacillus sp.SBS-4S:Cloning and characterization of the lipase[J].Journal of Bioscience and Bioengineering, 2011, 111(3):272-278.
[21] KUMAR S, STECHER G, LI M, et al.MEGA X:Molecular evolutionary genetics analysis across computing platforms[J].Molecular Biology and Evolution, 2018, 35(6):1 547-1 549.
[22] 克曌地. 金属离子与Candidasp.99-125脂肪酶相互作用及对水解汉麻油的影响[D].郑州:郑州大学,2017.
KE Z D.Effect of metal ions on the enzymatic hydrolysis of hemp seed oilby lipase Candida sp.99-125[D].Zhengzhou:Zhengzhou University, 2017.
[23] 祖昕,杜凯,曹方,等.有机溶剂处理提高固定化脂肪酶活性及稳定性[J].食品与生物技术学报,2016,35(1):89-94.
ZU X, DU K, CAO F, et al.Treatment of organic solvents to improve activity and stability of immobilized lipases[J].Journal of Food Science and Biotechnology, 2016,35(1):89-94.
