Bioinformatics and Expression Analysis of PksN in Bacillus velezensis Polyketide Synthase

doi:10.7668/hbnxb.20195156

Abstract

Abstract:

To explore the physicochemical properties,structure,and expression of the polyketide synthase PksN during polyketones biosynthesis in Bacillus velezensis DJ1 strain.It employed bioinformatics and molecular docking techniques to analyze the homologous sequences,physicochemical properties,protein structure,ligand-receptor binding sites,small molecule compounds,and protein-protein interaction networks of PksN.Additionally,Real-time Quantitative PCR was used to measure the expression of PksN at different time points during fermentation of Bacillus velezensis DJ1.The results showed that PksN belongs to the PksD family and shares 100% similarity with PksN gene of Bacillus.It contained 55 open reading frames(ORFs)and five conserved domains,including the KR domain.The protein consisted of 2 051 amino acids,with a molecular weight of 226.05 ku and an isoelectric point of 5.59.The molecular formula was C₁₀₀₂₈H₁₅₇₁₉N₂₇₅₃O₃₀₅₀S₇₅.PksN was an unstable protein located in the cytoplasmic membrane,and its secondary structure was predominantly α-helix.The tertiary structure of PksN was also predicted.Molecular docking results revealed that PksN can utilize acetyl-CoA,propionyl-CoA,isovaleryl-CoA,malonyl-CoA,methylmalonyl-CoA,and ethylmalonyl-CoA as substrates for polyketide biosynthesis.The top ten interacting proteins are all associated with polyketide biosynthesis.Real-time Quantitative PCR results indicated that the expression of PksN in Bacillus velezensis DJ1 peaked at 48 hours of fermentation,identifying the optimal time point for the subsequent extraction of polyketides.

Key words: Bacillus velezensis, Polyketide synthase, Bioinformatics, Structural prediction, Molecular docking, Gene expression

CLC Number:

Q936微生物生物化学

LIU Wei, YAN Gengxuan, HU Jihua, TIAN Yuan, YU Chong, MAO Xue, ZHANG Shumei. Bioinformatics and Expression Analysis of PksN in Bacillus velezensis Polyketide Synthase[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(S1): 292-299. doi: 10.7668/hbnxb.20195156.

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Figures/Tables 9

Tab.1 Real-time PCR primer sequence

基因Gene	引物序列(5'-3') Primer sequence(5'-3')
16S	GCTTCGGCTACCACTTACAGAT
	TCAGGTCGGCTACGCATCG
PksN	ATCGCCGATTTTGATCCTGAGTTC
	ATGTCCTCGTATATCCCGCATCC

Fig.1 PksN phosphorylation site prediction and hydrophobicity analysis results

Fig.2 PksN secondary structure

Fig.3 PksN three-dimensional structure

Tab.2 Binding scores between PksN and small molecules

小分子Micromolecule	得分Score
乙酰辅酶A Acetyl-CoA	-12.118 781 1
丙酰辅酶A Propionyl-CoA	-11.694 555 3
异戊酰辅酶A Isovaleryl-CoA	-11.775 791 2
丙二酰辅酶A Malonyl-CoA	-12.488 548 3
甲基丙二酰辅酶A Methylmalonyl-CoA	-12.189 192 8
乙基丙二酰辅酶A Ethylmalonyl-CoA	-13.497 603 4
甲酸Formic acid	-3.180 979 0
乙酸Acetic acid	-3.472 922 3
丙酸Propionic acid	-3.856 440 3
丁酸Butyric acid	-4.132 806 3

Fig.4 Binding patterns 1 between PksN and small molecules A.Acetyl CoA;B.Propionyl-CoA;C.Isovaleryl CoA;D.Malonyl-CoA; E.Methylmalonyl-CoA;F.Ethyl malonyl-CoA.Polar.Polar;Acidic.Acidic;Basic.Basic;Greasy.Hydrophobic;Proximity contour.Proximity contour;Sidechain acceptor.Sidechain acceptor;Sidechain.Denotes sidechain donor;Backbone acceptor.Backbone acceptor;Backbone.Backbone donor;Ligand exposure.Ligand exposure;Receptor exposure.Receptor exposure;Solvent residue.Solvent residue;Metal complex.Metal complex;Solvent contact.Solvent contact;Metal/ion contact.Metal/ion contact;Arene-arene.Arene-arene interaction;Arene-H.Arene-hydrogen interaction;Arene-cation.Arene-cation interaction.The same as Fig.5.

Fig.5 Binding patterns 2 between PksN and small molecules A.Formic acid;B.Acetic acid;C.Propionic acid;D.Butyric acid.

Fig.6 PksN protein interaction network

Fig.7 Relative expression of PksN at different fermentation times

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