Synthetic Gene DataBase
 

Synthetic Gene 196


 
  Welcome, Guest!

Field NameNatural GeneSynthetic Gene
SGDB Gene ID182196
GenBank AccessionAF217189DQ289491
GenBank GI745355482880264
Gene NameepoAepoA(SYN)
Gene Length (bp)42664326
SpeciesPolyangium cellulosum (Sorangium cellulosum)Escherichia coli
StrainsSMP44K207-3
CDSgtggcggatcgtcccatcgagcgcgcagccgaagatccgattgcgatcgtcggagcgggc
tgccgtctgcccggtggcgtgatcgatctgagcgggttctggacgctcctcgagggctcg
cgcgacaccgtcgggcaagtccccgccgaacgctgggatgcagcagcgtggtttgatccc
gacctcgatgccccggggaagacgcccgttacgcgcgcatctttcctgagcgacgtagcc
tgcttcgacgcctccttcttcggcatctcgcctcgcgaagcgctgcggatggaccctgca
catcgactcttgctggaggtgtgctgggaggcgctggagaacgccgcgatcgctccatcg
gcgctcgtcggtacggaaacgggagtgttcatcgggatcggcccgtccgaatatgaggcc
gcgctgccgcgagcgacggcgtccgcagagatcgacgctcatggcgggctggggacgatg
cccagcgtcggagcgggccgaatctcgtatgtcctcgggctgcgagggccgtgtgtcgcg
gtggatacggcctattcgtcctcgctcgtggccgttcatctggcctgtcagagcttgcgc
tccggggaatgctccacggccctggctggtggggtatcgctgatgttgtcgccgagcacc
ctcgtgtggctctcgaagacccgcgcgctggccacggacggtcgctgcaaggcgttttcg
gcggaggccgatgggttcggacgaggcgaagggtgcgccgtcgtggtcctcaagcggctc
agtggagcccgcgcggacggcgaccggatattggcggtgattcgaggatccgcgatcaat
cacgacggagcgagcagcggtctgaccgtgccgaacgggagctcccaagaaatcgtgctg
aaacgggccctggcggacgcaggctgcgccgcgtcttcggtgggttatgtcgaggcacac
ggcacgggcacgacgcttggtgaccccatcgaaatccaagctctgaatgcggtatacggc
ctcgggcgagacgtcgccacgccgctgctgatcgggtcggtgaagaccaaccttggccat
cctgagtatgcgtcggggatcactgggctgctgaaggtcgtcttgtcccttcagcacggg
cagattcctgcgcacctccacgcgcaggcgctgaacccccggatctcatggggtgatctt
cggctgaccgtcacgcgcgcccggacaccgtggccggactggaatacgccgcgacgggcg
ggggtgagctcgttcggcatgagcgggaccaacgcgcacgtggtgctggaagaggcgccg
gcggcgacgtgcacaccgccggcgccggagcggccggcagagctgctggtgctgtcggca
aggaccgcggcagccttggatgcacacgcggcgcggctgcgcgaccatctggagacctac
ccttcgcagtgtctgggcgatgtggcgttcagtctggcgacgacgcgcagcgcgatggag
caccggctcgcggtggcggcgacgtcgagcgaggggctgcgggcagccctggacgctgcg
gcgcagggacagacgccgcccggtgtggtgcgcggtatcgccgattcctcacgcggcaag
ctcgcctttctcttcaccggacagggggcgcagacgctgggcatgggccgtgggctgtat
gatgtatggcccgcgttccgcgaggcgttcgacctgtgcgtgaggctgttcaaccaggag
ctcgaccggccgctccgcgaggtgatgtgggccgaaccggccagcgtcgacgccgcgctg
ctcgaccagacagcctttacccagccggcgctgttcaccttcgagtatgcgctcgccgcg
ctgtggcggtcgtggggcgtagagccggagttggtcgctggccatagcatcggtgagctg
gtggctgcctgcgtggcgggcgtgttctcgcttgaggacgcggtgttcctggtggctgcg
cgcgggcgcctgatgcaggcgctgccggccggcggggcgatggtgtcgatcgcggcgccg
gaggccgatgtggctgctgcggtggcgccgcacgcagcgtcggtgtcgatcgccgcggtc
aacggtccggaccaggtggtcatcgcgggcgccgggcaacccgtgcatgcgatcgcggcg
gcgatggccgcgcgcggggcgcgaaccaaggcgctccacgtctcgcatgcgttccactca
ccgctcatggccccgatgctggaggcgttcgggcgtgtggccgagtcggtgagctaccgg
cggccgtcgatcgtcctggtcagcaatctgagcgggaaggctggcacagacgaggtgagc
tcgccgggctattgggtgcgccacgcgcgagaggtggtgcgcttcgcggatggagtgaag
gcgctgcacgcggccggtgcgggcaccttcgtcgaggtcggtccgaaatcgacgctgctc
ggcctggtgcctgcctgcctgccggacgcccggccggcgctgctcgcatcgtcgcgcgct
gggcgtgacgagccagcgaccgtgctcgaggcgctcggcgggctctgggccgtcggtggc
ctggtctcctgggccggcctcttcccctcaggggggcggcgggtgccgctgcccacgtac
ccttggcagcgcgagcgctactggatcgacacgaaagccgacgacgcggcgcgtggcgac
cgccgtgctccgggagcgggtcacgacgaggtcgagaaggggggcgcggtgcgcggcggc
gaccggcgcagcgctcggctcgaccatccgccgcccgagagcggacgccgggagaaggtc
gaggccgccggcgaccgtccgttccggctcgagatcgatgagccaggcgtgctcgatcgc
ctggtgcttcgggtcacggagcggcgcgcccctggtcttggcgaggtcgagatcgccgtc
gacgcggcggggctcagcttcaatgatgtccagctcgcgctgggcatggtgcccgacgac
ctgccgggaaagcccaaccctccgctgctgctcggaggcgagtgcgccgggcgcatcgtc
gccgtgggcgagggcgtgaacggccttgtggtgggccaaccggtcatcgccctttcggcg
ggagcgtttgctacccacgtcaccacgtcggctgcgctggtgctgcctcggcctcaggcg
ctctcggcgaccgaggcggccgccatgcccgtcgcgtacctgacggcatggtacgcgctc
gacggaatagcccgccttcagccgggggagcgggtgctgatccacgcggcgaccggcggg
gtcggtctcgccgcggtgcagtgggcgcagcacgtgggagccgaggtccatgcgacggcc
ggcacgcccgagaagcgcgcctacctggagtcgctgggcgtgcggtatgtgagcgattcc
cgctcggaccggttcgtcgccgacgtgcgcgcgtggacgggcggcgagggagtagacgtc
gtgctcaactcgctttcgggcgagctgatcgacaagagtttcaatctcctgcgatcgcac
ggccggtttgtggagctcggcaagcgcgactgttacgcggataaccagctcgggctgcgg
ccgttcctgcgcaatctctccttctcgctggtggatctccgggggatgatgctcgagcgg
ccggcgcgggtccgtgcgctcttcgaggagctcctcggcctgatcgcggcaggcgtgttc
acccctccccccatcgcgacgctcccgatcgctcgtgtcgccgatgcgttccggagcatg
gcgcaggcgcagcatcttgggaagctcgtactcacgctgggtgacccggaggtccagatc
cgtattccgacccacgcaggcgccggcccgtccaccggggatcgggatctgctcgacagg
ctcgcgtcagctgcgccggccgcgcgcgcggcggcgctggaggcgttcctccgtacgcag
gtctcgcaggtgctgcgcacgcccgaaatcaaggtcggcgcggaggcgctgttcacccgc
ctcggcatggactcgctcatggccgtggagctgcgcaatcgtatcgaggcgagcctcaag
ctgaagctgtcgacgacgttcctgtccacgtcccccaatatcgccttgttgacccaaaac
ctgttggatgctctcgccacagctctctccttggagcgggtggcggcggagaacctacgg
gcaggcgtgcaaagcgacttcgtctcatcgggcgcagatcaagactgggaaatcattgcc
ctatga
atggccgaccgcccgatcgaacgtgcagcggaggatccaattgcgattgtaggcgcgggc
tgccgcctgccgggcggcgtgattgacctctcgggcttctggacgctgttagaaggctcc
cgcgacaccgtcggtcaagtgccagcggagcggtgggatgctgcggcgtggttcgatccg
gatctggatgcacctggcaaaacaccagtgacccgcgccagctttttaagcgatgtcgcc
tgcttcgatgcctcttttttcgggatcagtccgcgcgaagcccttcgcatggatccggcc
caccggctgctgctggaagtgtgctgggaagcattggaaaacgcagctattgccccgtcg
gccctggttggcacggaaactggcgtctttattggcatcggtccaagcgaatatgaagcg
gcactgcctagggctactgccagcgcagaaattgatgctcacggcggcctgggcacgatg
ccttcagttggtgcaggtcgtatttcatacgtcctgggccttcgtggtccgtgtgtggcg
gtggacaccgcatatagttctagcttagtcgcagtacacctggcgtgtcagtcgttacgt
tccggcgaatgctcgaccgcgcttgcaggtggggtcagccttatgctgtccccgagcact
ttagtctggttgagcaagacacgtgcgttggcaaccgacggtcgctgcaaagccttcagc
gcggaggccgatgggtttggtcgtggcgaaggttgcgcagtggtcgtgctgaagcgtttg
tccggcgcacgtgcggatggggaccgcatcctcgcagttatccgcggctcggccatcaac
catgatggtgccagctccggtctcactgttccgaacggttcttcacaggaaattgtactg
aaacgcgccttagccgatgctggttgcgccgcatcttccgtggggtacgtcgaagctcat
gggacgggtactaccttaggcgatccgattgaaattcaggcgctcaatgccgtctacggc
ctgggtcgggatgtcgcgacccctttgctgatcgggtcggtcaagactaacctcggccat
ccagagtatgcctccgggatcactggtctgctgaaggttgtgttgtccttgcagcacggt
caaattccggcgcacctccatgctcaggcgttaaatccgcgcattagctggggcgatctg
cgtctgaccgttacccgtgctcggaccccgtggcctgactggaacacgcctcgccgcgcg
ggcgtctcctcgtttggcatgagtggtaccaatgcccacgttgttctggaggaagcccca
gcagcaacgtgcaccccgccagccccagaacgtccagccgaattgttagtgctgtctgcg
cgtaccgctgccgctctggacgcacatgcggcccgtttgcgcgaccatttagaaacatac
ccgtcacaatgtttaggtgacgttgccttctcgctggcgactacccgtagtgcgatggaa
catcgcctggcggtggccgctacgtcctcggagggtctgcgtgcggccttagacgccgca
gctcagggtcagaccccgccgggtgttgtccgtggtatcgcagactcgtctcgcggcaaa
ctggcttttctgtttactggccagggtgcccagacgctcggcatgggccggggcctgtac
gatgtttggcctgcttttcgcgaagcgtttgatttgtgtgtgcgcctgtttaaccaagaa
ctggatcgtccgctgcgtgaagtaatgtgggcagaaccagcatcagtagatgccgcactt
ttagaccagacagcttttacacagccagcgctttttacgtttgagtatgctctggctgca
ctgtggagatcttggggcgtagaaccagaactggtggccggtcactcgattggcgaactg
gtggcggcgtgcgttgcgggtgtgttcagtttggaggacgccgtgttcctggtcgcggca
cgcggtcgtctcatgcaggcgctgcctgctggtggtgcaatggtgtctattgcggcgcca
gaagcggacgtcgcggcggcggtcgcgcctcatgccgcatcagtaagtatcgcggctgtt
aatggcccagaccaagtggtaatcgcgggcgcagggcagccggtgcatgcgatcgccgct
gcaatggcggcgcgcggtgcccggaccaaagcgcttcacgtgagccacgcgttccacagt
ccactgatggcaccgatgttagaagcgtttggccgcgttgctgaatccgtaagttatcgt
cgtccgagcatcgtactcgttagtaatctgagcggcaaagcagggacagatgaagtatcc
agccctggctattgggtgcgtcatgctcgggaggttgtgcgtttcgcagatggcgtgaaa
gcgctccatgccgcaggtgcaggcacgtttgttgaagtgggtccgaagtctactcttttg
ggtttagttccggcgtgtttgccagacgctcgtccggcgcttctggcaagttctcgtgcc
gggcgcgatgaaccagccactgttctggaagctctggggggtctgtgggccgttggtggt
cttgtatcgtgggcaggtctgtttccgagtggcggtcgccgcgtgcctctgccgacgtat
ccgtggcaacgtgagcgttactggctgcagaccaaggcggatgacgcagcgcgtggtgat
cggcgagcaccgggtgcgggccatgacgaagtcgaaaaaggcggggcggtcagaggtggg
gatcgccgcagcgcccgtttggatcatccaccgccagagagcggacgccgtgaaaaggtg
gaggcagcgggcgaccgtccgtttcgtttggagattgatgagcctggcgtgctggaccgg
ctcgttctgcgtgttacggagcgtcgcgcaccgggcttaggtgaggtggaaattgctgta
gatgcggcaggtctgagttttaacgacgtgcagctggctctgggtatggttccggatgat
ctgccgggtaaaccgaatccgccgctgctgttaggcggggaatgtgccggccgcattgtg
gcggttggggaaggcgtaaatggtctggttgtaggtcagccggtgattgcactgagcgct
ggtgctttcgcaacccatgtcaccacgtcagccgccctggtgctgccacgccctcaggcg
ctgtccgcgaccgaggccgcagctatgccagtggcatatctcaccgcgtggtatgctctg
gatggcattgcccgccttcaacctggcgagcgcgtgctgatccatgcggccacgggtggc
gttggcctggcggcagtacagtgggcccagcacgtcggggccgaagttcacgctactgcg
ggtacgccagagaaacgcgcttaccttgaaagcctcggggttcgttacgtttcagattct
cgcagcgaccgctttgtagcagatgtgcgcgcctggaccggcggcgaaggcgttgatgtc
gttctgaactctctgtcaggtgaactgattgataagtcattcaacttactgcggtctcat
ggtcgttttgtcgaactcggcaaacgcgattgttatgctgataatcagctcggccttcgc
cctttcctgcgtaacctttcattttctttggttgatctgcgcggcatgatgctggaacgc
ccggcacgtgtgcgtgccttgtttgaggagctgctgggtttaattgccgctggtgtgttc
accccgccgccgatcgccacgcttcctattgctcgcgtggcggacgccttccgttcgatg
gcgcaagcacagcatttaggcaaactcgtactgaccctaggggatccggaggtccaaatc
cgtattccgacacacgcgggggccggtccgtctaccggcgaccgggacctgctggatcgt
cttgcgagtgctgcaccggcggctcgtgcggcggccttagaagcttttttgcgcacccag
gtgtcgcaagtgctgcgcacacctgaaattaaagtaggggctgaagctttgttcacacgg
ctgggtatggattccctgatggcagtggaacttcgtaatcgtattgaggcgagcttgaag
ctgaaattatctacaaccttccttagcacgagcccgaacatcgccctgctgacccaaaac
ttgttggatgcactctctagtgcattaagtttggaacgtgttgccgcggagaacctgcgc
gcgggcgtccaatccgactttgtgtcgtcaggggccgatcaggattgggaaatcattgct
ctggggaattgtagctccgtcgacaagcttgcggccgcactcgaacaccaccaccaccac
cactga
5' End
3' End
NotesNo accession number for the natural gene was mentioned in the paper, so the record AF217189 was used.
Expression VectorNApET22b
Assay MethodsNASDS-PAGE
ResultsPrevious attempts to expressed epoA in E. coli were unsuccessful (Chen et al. 2001, Chem. Biol.; O'Connor et al. 2002, Biochemistry)Coexpression of the E. coli chaperone proteins GroES/EL and trigger factor resulted in a large increase in the level of soluble EpoA protein.
Protein Functiontype I polyketide synthase
Recoding PurposeTo improve expression
Synthesized ByAuthors
Recoding MethodE.coli preferred codons were used for all amino acids. The designing of epoA gene was done by Ralph
Reid with DNAWorks. Amino acid subtitutions occurs to the following sites: I889L; D890Q; A1386S;
T1387S.
Publication Author(s)Mutka SC, Carney JR, Liu Y, Kennedy J.
Corresponding AuthorJonathan Kennedy
Corresponding AddressKosan Biosciences, Inc., 3832 Bay Center Place, Hayward, California 94545.
Publication Year2006
Publication TitleHeterologous Production of Epothilone C and D in Escherichia coli.
AbstractThe epothilones are a family of polyketide natural products that show a high potential as anticancer drugs. They are synthesized by the action of a hybrid nonribosomal peptide synthetase/polyketide synthase in the myxobacterium Sorangium cellulosum. In this work, the genes encoding the entire cluster,epoA, epoB, epoC, epoD, epoE, and epoF, were redesigned and synthesized to allow for expression in Escherichia coli. The expression of the largest of the proteins, EpoD, also required the protein be separated into two polypeptides with compatible module linkers. Using a combination of lowered temperature, chaperone coexpression, and alternative promoters, we succeeded in producing a soluble protein from all genes in the epothilone cluster. The entire synthetic epothilone cluster was then expressed in a strain of E. coli modified to enable polyketide biosynthesis, resulting in the production of epothilones C and D. Furthermore, feeding a thioester of the normal substrate for EpoD to cells expressing the epoD, epoE, and epoF genes also led to the production of epothilones C and D. The design of the synthetic epothilone genes together with E. coli expression provides the ideal platform for both the biochemical investigation of the epothilone PKS and the generation of novel biosynthetic epothilone analogues.
JournalBiochemistry. 2006 Jan 31;45(4):1321-30.. 45(4): 1321-30.
SummaryA cluster of epothilone genes were fully codon-optimized towards E. coli codon preference (with DNAWorks). It appeared that the synthetic epothilone genes were successfully expressed to a detectable level.
Comments
Discussion http://www.evolvingcode.net/forum/viewtopic.php?t=505
PubMed ID16430229
Submitter NameWu, Gang
Submitter AddressDepartment of Biological Sciences, University of Maryland Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250 USA
Entry ConfirmationNo
 
 

Copyright 2004 the Freeland Bioinformatics Lab, All Rights Reserved. | Contact Us | About this site