CN107974484A - Chalcone synthase modeling method in phloretin biosynthetic process - Google Patents
Chalcone synthase modeling method in phloretin biosynthetic process Download PDFInfo
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Abstract
The present invention relates to the model building method of chalcone synthase in phloretin biosynthetic process, step are as follows：1) homologous modeling；2) molecular docking：Substrate para hydroxybenzene propionic acid is docked to inside the active pocket of chalcone synthase, the molecular docking flow includes ligand and acceptor preparation, Grid generations, molecular docking, docking interpretation of result and PyMOL handling result file sub-steps；3) EbCHS catalytic mechanisms are predicted.The main innovation point of the present invention is as follows：1) based on the chalcone synthase three-dimensional structure of a variety of source of species, structure is suitable for the chalcone synthase model for catalyzing and synthesizing phloretin；2) the specific binding substrate of the chalcone synthase model is para hydroxybenzene propionic acid；3) the model docking of chalcone synthase and substrate para hydroxybenzene propionic acid small molecule is completed；4) the chalcone synthase model established is analyzed, and proposes corresponding Reconstruc-tion policy, foundation is provided to obtain efficient chalcone synthase.
Description
Technical field
The present invention relates to the model building method of chalcone synthase in phloretin biosynthetic process, belongs to synthesis life
Thing and field of bioinformatics.
Background technology
Phloretin is distributed mainly on the pericarp and root skin of the rich fruits such as apple, pears, and phloretin can suppress the mistake of sebaceous glands
Degree secretion, secretes vigorous type acne, moreover it is possible to suppress melanocyte activity, have necessarily to various skin splash for treating
Mitigation, phloretin is developed by foreign study in recent years is used as new type natural skin-whitening agents.When itself and ursin
And/or kojic acid can greatly improve inhibiting rate of the product to tyrosinase, inhibiting rate is reached 100% when being compounded.The whole world
20 tons are no less than per annual consumption, demand with the increase of 20% speed, has the larger market demand every year.
At present, the technique of existing extraction phloretin is mostly chemical method, there is sour water solution, enzymolysis and directly extraction.The method
There are the discharge of chemical waste fluid, chemical contamination are serious.Present inventor develops a kind of biological method synthesis phloretin,
Cheap substrate para hydroxybenzene propionic acid is added, by 4 coumarate CoA-ligase of heterogenous expression (4CL) and chalcone synthase
(EbCHS) phloretin is catalyzed and synthesized, this method output capacity is high, can continuous production.The problem of presently, there are is, in route of synthesis
Chalcone synthase catalytic efficiency it is limited, it is relatively low to the catalytic conversion of substrate, about 30%.
Meanwhile the modeling of enzyme molecule mainly passes through SWISS-MODEL, I-TASSER, RosettaCM in the prior art
Model Deng the homologous modeling of progress or from the beginning, using softwares such as AutoDock, DOCK6 or RosettaDock with molecular docking
Suitable substrate small molecule is docked in the active pocket of enzyme by means.Then the Candidate Residues to Enzyme-Substrate complex are passed through
Simulation random mutation optimization is carried out, then according to the size of mutant enzyme and corresponding substrate specificity complementarity and combination energy, sieve
The potential specific mutant of raising is selected, in order to improve the reactivity of enzyme, on the basis of specificity transformation result of calculation
On, it can suitably increase the scope of amino acid screening, such as increase the amino acid of substrate access way and enzyme surface, evolve not
Conservative amino acid, coevolution act on stronger amino acid etc., and the fixed point and saturation mutation then tested are improved with reaching
The purpose of activity of enzyme reaction.The present invention is exactly limited for the catalytic efficiency of chalcone synthase, to the catalytic conversion of substrate
Relatively low technical problem, there is provided a kind of chalcone synthase modeling and remodeling method.
The content of the invention
It is an object of the invention to solve the deficiencies in the prior art, there is provided Cha Er in a kind of phloretin biosynthetic process
Ketone synzyme modeling method.Main innovation point of the present invention is the model for completing chalcone synthase in phloretin building-up process
Structure and its molecular docking with substrate para hydroxybenzene propionic acid, and corresponding remodeling method is proposed according to model, to obtain higher
The chalcone synthase for phloretin synthesis of effect provides foundation, and synthesizing phloretin for Microbial cell factories lays the foundation.
The technical solution adopted by the present invention to solve the technical problems is：
Chalcone synthase modeling method in phloretin biosynthetic process, the method step are as follows：
1) homologous modeling：The homologous modeling procedure includes homologous templates search, sequence alignment and the life of Grishin files
Sub-step is completed into, structure thread file generateds and homologous modeling；
2) molecular docking：Substrate para hydroxybenzene propionic acid is docked to inside the active pocket of chalcone synthase, described point
Son docking flow includes ligand and acceptor preparation, Grid generations, molecular docking, docking interpretation of result and PyMOL handling results text
Part sub-step；
3) EbCHS catalytic mechanisms are predicted：According to step 1) and the modeling result and molecule of the 2) chalcone synthase of structure
Docking is predicted catalytic mechanism and key amino acid as a result, analyzed using Chimera protein three-dimensional structure
And provide certain mutation suggestion.
Preferably, homologous modeling concrete operations are as follows in step 1)：Homologous templates search for when, using EbCHS protein sequences as
Index, is retrieved in PDB databases, chooses candidate template of higher preceding 11 protein sequences of similarity as modeling, most
Sequence similarity and crystallization resolution ratio are considered eventually, are determined same as this using freesia chalcone synthase crystal structure 4wum
4wum, is first translated to Protein primary amino acid sequence, then by template sequence and chalcone synthase by the template of source modeling
Sequence is compared, and generates Grishin formatted files, according to result above, produces structure thread files, further according to
Thread files carry out homologous modeling.
Preferably, step 2) molecular docking carries out molecular docking using software AUTODOCK to EbCHS, substrate to hydroxyl
Base benzenpropanoic acid is docked to inside the active pocket of enzyme, and concrete operations are as follows：Ligand file and acceptor file are got out first, point
It is other that it is pre-processed, then with adt generation docking box used, docking order is carried out, docking result is divided
Analysis, recycles PyMOL handling result files, finds the receptor-ligand combinations for meeting pattern.
Preferably, the pretreatment specific method is：Ligand and acceptor molecule are pre-processed using adt softwares.
Ligand pre-processes：
Ligand ＞ Input ＞ Open ＞ HPA.pdb open small molecule
It is pdbqt files that Ligand ＞ Output ＞ Save as PDBQT ＞ HPA.pdbqt, which format small molecule,
Edit ＞ Delete ＞ Delete all molecular empty adt screens
Acceptor pre-processes：
Grid ＞ Macromolecular ＞ Open ＞ EbCHS.pdb open receptor protein
It is pdbqt files that Save as PDBQT ＞ EbCHS.pdbqt, which format acceptor,.
The species used in 1 present invention of table
| Species | Latin name | Gene | Remarks |
| Fleabane flower | Erigeron breviscapus | EbCHS |
Homologous modeling candidate template in 2 present invention of table
The beneficial effects of the invention are as follows：The present invention synthesizes living things catalysis fleabane flower (Erigeron during phloretin
Breviscapus) chalcone synthase (chalcone synthase, EbCHS) in source carries out homologous modeling, molecule pair
Connect, and the model to building up is analyzed, and proposes corresponding modification scheme, for obtain efficient chalcone synthase provide according to
According to.
The main innovation point of the present invention is as follows：
1) based on the chalcone synthase three-dimensional structure of a variety of source of species, structure is suitable for catalyzing and synthesizing phloretin
Chalcone synthase model；
2) the specific binding substrate of the chalcone synthase model is para hydroxybenzene propionic acid；
3) the model docking of chalcone synthase and substrate para hydroxybenzene propionic acid small molecule is completed；
4) the chalcone synthase model established is analyzed, and proposes corresponding Reconstruc-tion policy, to obtain efficiently
Chalcone synthase provide foundation.
Brief description of the drawings
It is the phloretin synthetic route chart described in this project to illustrate Fig. 1
Fig. 2 is the EbCHS protein sequences described in this project
Fig. 3 is the EbCHS-4wum sequence alignment results described in this project
Fig. 4 is the Grishin file formats described in this project implementation process
Fig. 5 is EbCHS protein structures and activated centre in this project implementation process
Fig. 6 is homologous modeling and molecular docking flow chart in this project implementation process
Embodiment
Below by specific embodiment, and with reference to attached drawing, technical scheme is described in further detail.
1 homologous modeling of embodiment
The present invention uses the rosettaCM modules in software ROSETTA suite to carry out homologous modeling, modeling to EbCHS
Flow such as Fig. 6 a, are broadly divided into the following steps：
1) homologous templates are searched for：Indexed with the protein sequence (Fig. 2, annex one) of EbCHS for search, to PDB databases
(network address：http://www.rcsb.org/pdb/home/home.do) retrieved, 141 are obtained with certain similar
The protein structure of degree, takes first 11 candidate templates (table 2) as this homologous modeling.By to this 11 homologous templates
It is compared, finds in the protein structure of these same or similar functions, freesia chalcone synthase crystal structure 4wum
On the premise of higher similarity is possessed, the resolution ratio of crystal diffraction is also of a relatively high, because the A chain conducts of selection 4wum
The template of this simulation；
2) sequence alignment and Grishin file generateds.
First, using pdb2seq.py scripts, protein structure file 4wumA is translated amino acid sequence, is obtained
4wumA.fasta, order are as follows：
[user@localhost home]$pdb2seq.py 4wumA.pdb
Then EbCHS.fasta and 4wumA.fasta two sequences are handled, obtains EbCHS_4wumA.fasta
(two sequences are merged into a fasta file), afterwards using sequence alignment program clustalo to two sequences into
Row compares, and obtains pairwise comparison file EbCHS_4wumA.vie (Fig. 3), and order is as follows：
[user@localhost home]$clustalo-i EbCHS_4wumA.fasta-o EbCHS_4wumA.vie-
v-outfmt vie-threads 4--force
After EbCHS_4wumA.vie is obtained, the call format (network address with reference to rosettaCM to Grishin files：
https://www.rosettacommons.org/docs/latest/rosetta_basics/file_types/ Grishan-format-alignment), EbCHS_4wumA.vie files are converted to EbCHS.grishin files, see figure
4。
3) structure thread file generateds.
By first two steps, Multiple Sequence Alignment file EbCHS.grishin, formwork structure 4wumA.pdb and ammonia are obtained
Base acid sequence EbCHS.fasta, using these files as input, is ordered using partial_thread.linuxgccrelease
Order produces structure thread files：
[user@localhost home]$partial_thread.linuxgccrelease@thread_flags
Note：Thread_flags files are order configuration file, see annex two.
After order is finished, 4wumA_thread.pdb files are obtained, this file is using as the defeated of homologous modeling procedure
Enter file.
4) homologous modeling.
First, the energy and scoring functions for homologous modeling configure Parameter File rosettaCM_2014.xml, in detail letter
Breath is shown in annex four.
Then with EbCHS.fasta files, rosettaCM_2014.xml files and 4wumA_thread.pdb files
For input, homologous modelling command is run, it is as follows：
[user@localhost home]$mpirun-np 8 rosetta_scripts.linuxgccrelease@
model_flags
Note：Mode1_flgas files are modelling command configuration file, see annex three.
Openmpi is used to run modelling command at the same time using 8 kernels, altogether as multithreading running environment in this order
500 models are built, choose the highest result as this modeling that wherein scores.
2 molecular docking of embodiment
The present invention carries out molecular docking using software AUTODOCK to EbCHS, and substrate para hydroxybenzene propionic acid is docked to enzyme
Active pocket inside.Flow such as Fig. 6 b are docked, are broadly divided into the following steps：
1) ligand file and acceptor file prepare.
The ligand file of this time molecular docking is HPA.pdb, and acceptor file is EbCHS.pdb.First by adt softwares
Ligand and acceptor molecule are pre-processed.
Ligand pre-processes：
Ligand ＞ Input ＞ Open ＞ HPA.pdb open small molecule
It is pdbqt files that Ligand ＞ Output ＞ Save as PDBQT ＞ HPA.pdbqt, which format small molecule,
Edit ＞ Delete ＞ Delete all molecular empty adt screens
Acceptor pre-processes：
Grid ＞ Macromolecular ＞ Open ＞ EbCHS.pdb open receptor protein
It is pdbqt files that Save as PDBQT ＞ EbCHS.pdbqt, which format acceptor,
2) Grid is generated.
It is as follows using adt generation docking box used, detailed step after having handled ligand molecular and acceptor molecule：
Grid ＞ Macromolecular ＞ Open ＞ EbCHS.pdbqt open receptor protein
Grid ＞ Set Map Types ＞ Open Ligand ＞ HPA.pdbqt open small molecule file
Grid ＞ Grid box ＞ set grid center ＞ to set xyz and center ＞ File ＞ Close saving
current
Grid ＞ Out Put ＞ grid.gpf preserve grid Parameter File
[user@localhost home]$autogrid4-p grid.gpf-l grid.glg
The lattice point file of .map endings is generated, button d.gpf files are shown in annex five.
3) molecular docking.
Previous step is handled after obtaining lattice point Parameter File, this step sets docking parameter and then performs docking order：
Edit ＞ Delete ＞ Delete All Molecular empty adt screens
Docking ＞ Macromolecule ＞ Set Rigid Filename ＞ EbCHS.pdbqt
Docking ＞ Ligand ＞ Open ＞ HPA.pdbqt open docking ligand file
Docking ＞ Search Parameters ＞ Number of GA runs ＞ 20
Docking ＞ Output ＞ Lamarckian GA ＞ dock.dpf
[user@localhost home]$autodock4-p dock.dpf-l dock.dlg
Dock.dlg docking destination files are generated, dock.dpf files are shown in annex six.
4) interpretation of result is docked.
Analyze ＞ Docking ＞ Open ＞ dock.dlg open docking destination file
Analyze ＞ Macromolecuar ＞ Open ＞ EbCHS.pdbqt open acceptor file
Analyze ＞ Clusterings ＞ Show show sub-clustering information
5) PyMOL handling results file.
Docking destination file is handled using PyMOL, finds the receptor-ligand combinations for meeting binding pattern.
PyMOL ＞ open ＞ EbCHS.pdb open acceptor pdb
PyMOL ＞ open ＞ All files ＞ dock.dlg open docking result
File ＞ save molecule ＞ EbCHS-HPA.pdb preserve suitable receptor-ligand combinations file
Obtain molecular docking destination file EbCHS-HPA.pdb
Embodiment 3EbCHS catalytic mechanisms are predicted
According to the modeling result of above-mentioned steps and molecular docking as a result, being painted using Chimera to protein structure figure
System, and supposition analysis is carried out to catalytic mechanism and key amino acid and mutation suggestion.
1) key amino acid.After the binding pattern of analysis HPA and protein receptor, applicants have discovered that the binding domain of HPA
Mainly it is made of following amino acid：V101, T135, S136, G166, C167, E195I196, T197, F168, A198, V199,
T200, F218, I257, L266, F268, S339, S341, S342, G377, P378 etc..These amino acid are mainly formed with HPA
Van der Waals force, so as to stablize the conformation of HPA, wherein two key amino acids of C167, I196 phenolic hydroxyl group and carbonyl with HPA respectively
Base oxygen forms interaction of hydrogen bond, and binding pattern such as Fig. 4, wherein C167 may participate in the fracture of CoA derivatives S-C keys.
2) catalytic mechanism.The catalytic mechanism of HPA and the synthetic route of phloretin are as shown in Figure 1.According to the knot of molecular docking
Fruit finds that the c-terminus of HPA makes malonyl coenzyme A be easier attack HPA carbonyl carbons towards substrate binding pocket outside.
Two amino acid of C167, I196 is beneficial to the conformational stability of HPA, while the presence of C167 with the hydrogen bond that HPA is formed, and coacetylase is spread out
The S-C keys more easy fracture of biology, the presence of wherein H306 and C339 are easier to the transmission of electronics when HPA and COA reacts, make
C167 is changed into S-, it is easier to realizes the attack to carbonyl carbon.
3) mutation is suggested.The mutation suggestion of the application is concentrated mainly on two aspects：
A) accelerate the electron transmission or structure basic amino acid of C167, the S-C keys of malonyl COA are more easily broken off
And further enhance stability of the HPA in pocket.Amino acid mutation suggestion is：S341E/D, N339Q, F268E/Q.
B) increase chain carrier, accelerate the substrate of HPA and malonyl coenzyme A to pass in and out, be malonyl coenzyme A faster
With HPA react.Amino acid mutation suggestion is：F168A, L270A.
Annex one：EbCHS.fasta
＞ EbCHS
MASSIDIAAIREAQRAQGPATILAIGTATPSNCVYQADYPDYYFRITKSEHMVDLKEKFKRMCDKS
MIRKRyMHLTEEYLKENPSLCEYMAPSLDARQDVVVVEVPKLGKEAATKAIKEWGQPKSKITHLI
FCTTSGVDMPGADYQLTKLLGLRPSVKRFMMYQQGCFAGGTVLRLAKDLAENNKGARVLVVCS
EITAVTFRGPNDTHLDSLVGQALFGDGAAAVIVGSDPDLTTERPLFEMISAAQTILPDSEGAIDGHLR
EVGLTFHLLKDVPGLISKNIEKALTQAFSPLGISDWNSLFWIAHPGGPAILDQVELKLGLKEEKMRA
TRHVLSEYGNMSSACVLFIIDEMRKKSAEDGAATTGEGLDWGVLFGFGPGLTVETVVLHSLPTTT AIAT
Annex two：thread_flags
-database RosettaHome/main/database
-in：file：fasta Outputs/Alignments/EbCHS.fasta
-in：file：alignment Outputs/Alignments/EbCHS.Grishin
-in：file：template_pdb 4wumA.pdb
-ignore_unrecognized_res
Annex three：model_flags
-database RosettaHome/main/database
-in：file：fasta Outputs/Alignments/EbCHS.fasta
-parser：protocol Config/rosettaCM_2014.xml
-nstruct 500
-relax：rminimize_bond_angles
-relax：min_type lbfgs_armijo_nonmonotone
-relax：jump_move true
-relax：default_repeats 3
-relax：dualspace
-out：sumx_0
-default_max_cycles 200
-out：path：all./Outputs/Models
.overwrite
Annex four：rosettaCM_2014.xml
<ROSETTASCRIPTS>
<TASKOPERATIONS>
</TASKOPERATIONS>
<SCOREFXNS>
<Stage1 weights=score3 symmetric=0>
<Reweight scoretype=atom_pair_constraint weight=0.5/>
</stage1>
<Stage2 weights=score4_smooth_cart symmetric=0>
<Reweight scoretype=atom_pair_constraint weight=0.5/>
</stage2>
<Fullatom weights=talaris2013_cart symmetric=0>
<Reweight scoretype=atom_pair_constraint weight=0.5/>
</fullatom>
</SCOREFXNS>
<FILTERS>
</FILTERS>
<MOVERS>
<Hybridize name=hybridize stage1_scorefxn=stage1 stage2_scorefxn=
Stage2 fa_scorefxn=fullatom batch=1 stage1_increase_cycles=1.0 stage2_
Increase_cycles=1.0 linmin_only=0 realign_domains=0>
<Template pdb=" 4wumA_thread.pdb " cst_file=" AUTO " weight=1.000/>
</Hybridize>
<FastRelax name=" relax " scorefxn=talaris2013/>
</MOVERS>
<APPLY_TO_POSE>
</APPLY_TO_POSE>
<PROTOCOLS>
<Add mover=hybridize/>
<Add mover=relax/>
</PROTOCOLS>
<OUTPUT scorefxn=talaris2013/>
</ROSETTASCRIPTS>
Annex five：grid.gPf
Annex six：dock.dpf
Embodiment described above is a kind of preferable scheme of the present invention, and not the present invention is made in any form
Limitation, also have other variations and remodeling on the premise of without departing from the technical solution described in claim.
Claims (5)
1. chalcone synthase modeling method in phloretin biosynthetic process, it is characterised in that：The method step is as follows：
1) homologous modeling：The homologous modeling procedure includes homologous templates search, sequence alignment and Grishin file generateds, knot
Sub-step is completed in structure thread file generateds and homologous modeling；
2) molecular docking：Substrate para hydroxybenzene propionic acid is docked to inside the active pocket of chalcone synthase, the molecule pair
Connecing flow includes ligand and acceptor preparation, Grid generations, molecular docking, docking interpretation of result and PyMOL handling results file
Step；
3) EbCHS catalytic mechanisms are predicted：According to step 1) and 2) modeling result of the chalcone synthase of structure and molecular docking
As a result, being analyzed using Chimera protein three-dimensional structure, catalytic mechanism and key amino acid are predicted and provided
Certain mutation suggestion.
2. chalcone synthase modeling method in phloretin biosynthetic process according to claim 1, it is characterised in that：
Homologous modeling concrete operations are as follows in step 1)：When homologous templates are searched for, using EbCHS protein sequences as index, in PDB numbers
According to being retrieved in storehouse, the highest PDB templates of the similarity retrieved are freesia chalcone synthase crystal structure 4wum, first 4wum
Protein primary amino acid sequence is translated to, then template sequence and chalcone synthase sequence are compared, is generated
Grishin formatted files, according to result above, produce structure thread files, and homologous modeling is carried out further according to thread files.
3. chalcone synthase modeling method in phloretin biosynthetic process according to claim 2, it is characterised in that：
The amino acid sequence of the chalcone synthase genes comes from fleabane flower.
4. chalcone synthase modeling method in phloretin biosynthetic process according to claim 1, it is characterised in that：
Step 2) molecular docking carries out molecular docking using software AUTODOCK to EbCHS, and substrate para hydroxybenzene propionic acid is docked
To the active pocket of enzyme, concrete operations are as follows：Ligand file and acceptor file are got out first, it is located in advance respectively
Reason, then with adt generation docking box used, carries out docking order, docking result is analyzed, is recycled at PyMOL
Destination file is managed, finds the receptor-ligand combinations for meeting binding pattern.
5. chalcone synthase modeling method in phloretin biosynthetic process according to claim 4, it is characterised in that：
It is described pretreatment specific method be：
Ligand and acceptor molecule are pre-processed using adt softwares.
Ligand pre-processes：
Ligand ＞ Input ＞ Open ＞ HPA.pdb open small molecule
It is pdbqt files that Ligand ＞ Output ＞ Save as PDBQT ＞ HPA.pdbqt, which format small molecule,
Edit ＞ Delete ＞ Delete all molecular empty adt screens
Acceptor pre-processes：
Grid ＞ Macromolecular ＞ Open ＞ EbCHS.pdb open receptor protein
It is pdbqt files that Save as PDBQT ＞ EbCHS.pdbqt, which format acceptor,.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109913508A (en) * | 2018-06-05 | 2019-06-21 | 嘉兴欣贝莱生物科技有限公司 | A method of phloretin is synthesized using cyanobacteria |
| CN110343680A (en) * | 2019-08-01 | 2019-10-18 | 济南广盛源生物科技有限公司 | A kind of honeysuckle chalcone synthase mutant and application thereof |
| CN111748532A (en) * | 2020-05-25 | 2020-10-09 | 天津大学 | Application of novel p-coumaroyl-CoA ligase in biosynthesis of phloretin |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2016016826A1 (en) * | 2014-08-01 | 2016-02-04 | Andrew Patrick Dare | Methods and materials for manipulating phloridzin production |
| CN105765592A (en) * | 2013-09-27 | 2016-07-13 | 科德克希思公司 | Automated screening of enzyme variants |
-
2017
- 2017-11-10 CN CN201711111219.9A patent/CN107974484A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105765592A (en) * | 2013-09-27 | 2016-07-13 | 科德克希思公司 | Automated screening of enzyme variants |
| WO2016016826A1 (en) * | 2014-08-01 | 2016-02-04 | Andrew Patrick Dare | Methods and materials for manipulating phloridzin production |
Non-Patent Citations (3)
| Title |
|---|
| PRAVEEN AWASTHI ET AL.: "Homology modeling and docking study of chalcone synthase gene (CfCHS) from Coleus forskohlii", 《COGENT BIOLOGY》 * |
| 刘涛 等: "灯盏花查尔酮合成酶基因表达与灯盏乙素含量关系的研究", 《中国中药杂志》 * |
| 郁凯: "以计算机辅助分子设计方法改造谷氨酸脱羧酶的研究", 《中国博士学位论文全文数据库（工程科技I辑）》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109913508A (en) * | 2018-06-05 | 2019-06-21 | 嘉兴欣贝莱生物科技有限公司 | A method of phloretin is synthesized using cyanobacteria |
| CN110343680A (en) * | 2019-08-01 | 2019-10-18 | 济南广盛源生物科技有限公司 | A kind of honeysuckle chalcone synthase mutant and application thereof |
| CN110343680B (en) * | 2019-08-01 | 2020-03-17 | 济南广盛源生物科技有限公司 | Honeysuckle chalcone synthase mutant and application thereof |
| CN111748532A (en) * | 2020-05-25 | 2020-10-09 | 天津大学 | Application of novel p-coumaroyl-CoA ligase in biosynthesis of phloretin |
| CN111748532B (en) * | 2020-05-25 | 2022-04-15 | 天津大学 | Application of novel p-coumaroyl-CoA ligase in biosynthesis of phloretin |
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