Azino_00250 Azino_00250   Azino_00270 Azino_00270

Azino_00260 : CDS information

close this sectionLocation

Organism
StrainNRRL 2485 (=NBRC 13928)
Entry nameAzinomycin B
Contig
Start / Stop / Direction38,515 / 43,854 / + [in whole cluster]
38,515 / 43,854 / + [in contig]
Location38515..43854 [in whole cluster]
38515..43854 [in contig]
TypeCDS
Length5,340 bp (1,779 aa)
Click on the icon to see Genetic map.

close this sectionAnnotation

Category1.1 PKS
Productpolyketide synthase
Product (GenBank)Azi26
GeneaziB
Gene (GenBank)azi26
EC number
Keyword
  • iterative
  • 5-methyl-naphthoic acid
Note
Note (GenBank)
  • iterative type I PKS: KS-AT-DH-KR-ACP
Reference
ACC
PmId
[18635006] Characterization of the azinomycin B biosynthetic gene cluster revealing a different iterative type I polyketide synthase for naphthoate biosynthesis. (Chem Biol. , 2008)
[26731610] Polyketide Ring Expansion Mediated by a Thioesterase, Chain Elongation and Cyclization Domain, in Azinomycin Biosynthesis: Characterization of AziB and AziG. (Biochemistry. , 2016)
comment
[PMID: 18635006](2008)
Azinomycin B生合成遺伝子クラスターの報告。

aziB: Iterative type I PKS (KS-AT-DH-KR-ACP)

iterative decarboxylative condensationにより1つのacetyl-CoAと5つのmalonyl-CoAsからnascent linear hexaketideのアセンブリが行われるのに関与すると推測している。

aziBのみをhost S.albusにて発現させた株(AL1004)の培養上清は5-methyl-naphthoic acid(NPA)を産生したことから、bicyclic aromatic compound 5-methyl-NPAの生合成にiterative type I PKSとしてAziBが関与する事が明白であることを示した。

aziBのKR domainとDH domainをそれぞれinactiveにしたものをhost S.albusにて発現させた株を作製し、5-methyl-NPA形成のためにAziBのKR,DH domainは必須であることを確かめた。

buliding block 3-methoxy-5-methyl-NPAの生合成経路を確立するため、S.albusにaziB, B1, B2を導入した株を作製し、3-methoxy-5-methyl-NPAの産生を確認した。この結果から、配列解析で割り当てられたaziB1, B2の機能が確認され、azinomycin B骨格を組み立てるNRPS AziA1の基質として3-methoxy-5-methyl-NPAが強く支持された。

AziB, NcsB, ChlB1, MdpB, MchA, WcbR(control)のKR Domain配列でアライメント作製。
AziBのKR配列は触媒残基(K, S, Q, Y, N)を全て保存していたことから、keto基を還元する活性があると推測した。その一方、active site groove(溝)のH残基と特徴づけられたLDD motifが保存されていなかったので、AziBのKR domainはA1 type KR familyであり、もっぱらL-hydroxy構造を産出するためにketo基で機能すると推測された。

---
[PMID: 26731610](2016)
予想と異なりPKS(AziB)だけでは5-methyl naphthoate部分を形成することができず、2-methylbenzoic acidが形成された。thioesterase(AziG)が、AziBだけでは足りない２ラウンドの鎖伸長と環化を担う。
Related Reference
ACC
P87162
PmId
comment
BLAST id38%
Aspergillus terreus_atX
6-methylsalicylic acid synthase

6-methylsalicylic acid synthase_ATXのdehydratase(DH) domainの機能的検証。
DH domainはACPに繋がれたbeta-hydroxytriketide中間体のdehydrationに関連しないが、ACPから6- methylsalicylic acidを放出するためにthioester hydrolysisを触媒することが明らかになった。
よってDH domainからTH domain(Gly904 - Thr1048の145aa)へ改名。

構造解析報告のあるEryDH4とのalignmentから、H972XXXGXXXXP motif and Asp1129 が触媒に重要な残基であると提唱されている。著者らの既報で、ATX H972A mutantがbeta-hydroxy triketideのような産物をもたらさないことが確認されている。

close this sectionPKS/NRPS Module

A1 acetyl-CoA
malonyl-CoA
KS11..383
AT546..858
TH865..1017
KR1389..1582
ACP1666..1739

close this sectionSequence

selected fasta
>polyketide synthase [Azi26]
MAENVQNPPVEPLAVIGMSCRFAPDLDTPGRLWEFLRAGGSAVGEMPDRRWDPYVTDSRT
RDILRTTTRKGSFMRDIEGFDAEFFQITPREAEYIDPQQRIMLELAWEALCDAGLPPTSL
AGTDASVYVAANSNDYGRRLLEDLDRTGAWAVNGTTFYGIANRISYFLDAHGPSMAVDTA
CAGSLTALHVAGQALHRGETSVAIVGGINIMASPALVVALDAASATSPDGRSKSFDKAAD
GYGRGEGGGVVVLKRLSDAVRDGDPVHGLVLASGVFQDGRSDGMMAPNGSAQQRMLEEIY
RRSGIDPGTVQYVEAHGTGTQLGDAAEAQAIGNVFGPGRDGDNPLLIGTLKPNVGHVEAA
SGIAGVIKVLLGMRHGELPPSPHEEPDPGLGLEARGLRLVAEPTPWPRGEHGMRAGVSSY
GVGGSIAHAVLQQAPPRPDRTERPAAAATGRPQVFPLSAASEQGVRGLAGSVAAWLRAHP
ETALDDLAHTFTARRSHLSRRAAVVAGTTEELLGGLDALAGGEKSPAVALASASGFGDGG
AAGPAWVFSGHGAQWSGMGRELLTTEPVFAQVIDELAPVFSEELGWTPREAIEAGGPWTV
VRTQAMTFAMQVALAEVWSDLGLRPGAIIGHSVGEIAAAAVAGSLDRAEAARFACRRARA
LGKIAGRGAMAMVPMAFADVEQRVAGRDAVVAAIAASPLSTVVSGDTAAVEALLADLEAD
GIQARRVNTDVAFHSPHVQEILDEVRQAAAALRAGTPRVTLYSTALADPRSDAPREGEYW
ATNLADPVRFHQAVRAALDDGTRVFLEVSSHPVVAHSITETALDAGVPDAHVAITLRREQ
PEQRTVLANLARLHSLGTPVTWSYDGDLVDVPAVRWQHKPYWIFPDTAPEQGAGLGHDPQ
THTLIGARTTVASAPVQRVWQTELHMENRPYAQSHKVVGVETVPASVVLNSFITAATNEG
ERACGLRDIVFRIPLAAHPTRVVQVVLEQDKVRIASRIKRDQESGGVRDDEWLTHTTATV
VHEPEVGARPMEDPDVIRARCPVSWTWAKVDGIFRTMGVDGYTFPWVVEELLRGEDEQFS
TITVDHTPKLHPSSWTAVVDAALTASGVLVMDENSNVLRTCSHLESLSFVGPPPPRIHVH
TVRDPRTPDTISMTVADESGAVVCEARGLRYVKVQDIGSGAVGPRDLVHELAWEPVEVPA
DAPVPSQALVVGGAAGGPALVEALTARGVRARAVPDATAIGDASLTCADVVVVAPEALLP
GEAPEQAARRCAQLLVDAVQQVAAVPDERRRPRVWALTREVRAGATEAALAHAPLWGAGR
IVAGERPDLWGGVIDVAENAVPQQVASLIGALPHTEDVLSLDSEGVTAARLRQVARPAER
EPVDCRPDGTYLVTGGLGALGLEAARHLVEQGARRLVLIGRRGLPSRSRWDQVDDPAVAA
QIAEVVALEAAGATVRVLSLDISDAEATARALDPGALDMPPVRGIVHCAGVVSDALVEKT
GAANLDTTMGPKADGAMVLHRLFPAGTLDFFTMFSSCGQLARLTGQVSYASANSFLDALA
ALRRSRGETGTTSFAWAQWIGRGMGETTGRATILEAESRGLGGITVSEALRSWAYADRFA
LPYAAVMRVMPDHTLPVFSHLSVTDAGAQSADAGGVDWATVPAGELPELVLKVTHEQVAA
ELNLAVDDIAIDQPLLELGVDSVLTVALRVRLHRCFAVDLPPTILWSNPTVRALAEFLAA
EVGGATADAEETDPVAGLPAPQQGSGTAEQLDAVAAAAG
selected fasta
>polyketide synthase [Azi26]
TTGGCTGAGAATGTGCAGAACCCGCCCGTGGAGCCCCTCGCGGTGATCGGCATGAGCTGC
CGGTTCGCCCCCGACCTCGACACCCCCGGCCGGCTCTGGGAGTTCCTGCGCGCCGGGGGC
AGCGCGGTGGGCGAGATGCCCGACCGGCGCTGGGACCCGTACGTGACGGACAGCCGCACC
CGCGACATCCTGCGCACGACCACCCGCAAGGGTTCGTTCATGCGGGACATCGAGGGCTTC
GACGCCGAGTTCTTCCAGATCACGCCCCGCGAGGCCGAGTACATCGACCCGCAGCAGCGG
ATCATGCTCGAACTCGCCTGGGAAGCCCTGTGCGACGCCGGACTGCCGCCGACCTCGCTG
GCCGGCACCGACGCGAGCGTCTACGTGGCGGCGAACTCCAACGACTACGGCCGCCGCCTG
CTCGAAGACCTGGACCGCACGGGCGCCTGGGCCGTGAACGGCACCACCTTCTACGGCATC
GCCAACCGCATCTCGTACTTCCTGGACGCGCACGGCCCCAGCATGGCCGTGGACACCGCC
TGCGCCGGATCCCTGACCGCCCTGCACGTCGCGGGCCAGGCGCTGCACAGGGGCGAGACG
AGCGTGGCCATCGTCGGCGGCATCAACATCATGGCCTCGCCGGCGCTGGTCGTGGCCCTG
GACGCGGCCTCCGCGACCTCCCCGGACGGCCGCAGCAAGTCGTTCGACAAGGCGGCCGAC
GGCTACGGGCGCGGCGAGGGCGGCGGAGTCGTCGTGCTCAAGCGACTGTCCGACGCGGTG
CGCGACGGTGACCCGGTGCACGGGCTCGTCCTGGCCAGCGGTGTCTTCCAGGACGGCCGC
TCCGACGGCATGATGGCCCCCAACGGCAGTGCCCAGCAGCGCATGCTGGAAGAAATCTAC
CGGCGGTCCGGCATCGACCCCGGCACAGTGCAGTACGTCGAGGCGCACGGCACCGGCACC
CAGCTCGGTGACGCCGCGGAGGCCCAGGCCATCGGCAATGTCTTCGGCCCCGGCAGGGAC
GGCGACAACCCCCTGCTGATCGGGACGCTGAAGCCGAACGTGGGCCACGTCGAGGCAGCC
TCCGGCATCGCGGGCGTGATCAAGGTCCTGCTGGGGATGCGCCACGGCGAACTGCCGCCC
AGCCCGCACGAGGAGCCCGATCCCGGGCTCGGGCTGGAGGCCAGGGGACTGCGTCTGGTC
GCCGAGCCGACGCCGTGGCCGCGGGGCGAACACGGCATGCGCGCCGGTGTCTCCAGCTAC
GGCGTGGGCGGCTCGATCGCCCACGCCGTACTGCAGCAGGCCCCGCCGCGCCCGGACAGG
ACCGAGCGCCCGGCCGCCGCCGCCACGGGCCGCCCCCAGGTCTTCCCGCTGTCCGCCGCC
TCCGAGCAGGGCGTACGAGGACTGGCCGGCTCGGTCGCCGCGTGGCTGCGGGCACACCCC
GAGACCGCCCTGGACGATCTCGCCCACACCTTCACCGCCCGCCGCTCCCATCTCTCCCGC
CGGGCCGCCGTCGTGGCCGGCACCACGGAGGAACTGCTCGGCGGACTGGACGCGTTGGCC
GGCGGCGAGAAATCCCCCGCGGTGGCCCTGGCATCGGCGTCCGGGTTCGGTGACGGCGGC
GCGGCCGGACCCGCCTGGGTGTTCTCCGGGCACGGCGCGCAGTGGTCCGGCATGGGCCGG
GAACTGCTCACCACCGAGCCCGTGTTCGCCCAGGTCATCGACGAACTCGCCCCGGTCTTC
TCCGAAGAGCTCGGATGGACCCCGCGCGAGGCGATCGAGGCGGGCGGCCCGTGGACCGTC
GTCCGTACCCAGGCGATGACCTTCGCGATGCAGGTCGCCCTCGCCGAGGTGTGGTCGGAC
CTGGGCCTGCGTCCGGGCGCGATCATCGGGCACTCGGTCGGCGAGATCGCCGCCGCCGCG
GTGGCCGGGAGCCTGGACCGCGCAGAGGCGGCACGCTTCGCCTGCCGCCGCGCCCGCGCC
CTCGGAAAGATCGCCGGGCGTGGCGCGATGGCGATGGTGCCCATGGCGTTCGCCGACGTC
GAACAGCGGGTCGCCGGCCGGGACGCAGTGGTCGCGGCGATCGCCGCCAGCCCTCTGTCG
ACCGTGGTGTCGGGTGACACGGCCGCGGTCGAGGCCCTGCTCGCGGACCTGGAGGCCGAC
GGCATCCAGGCTCGCCGGGTCAACACCGACGTCGCCTTCCACAGCCCGCACGTCCAGGAG
ATCCTCGACGAGGTCAGGCAGGCCGCCGCCGCACTGCGGGCCGGAACACCCCGGGTGACG
CTGTACAGCACCGCCCTCGCGGACCCGCGCTCCGACGCTCCCCGTGAGGGCGAGTACTGG
GCCACCAACCTGGCCGACCCGGTCCGCTTCCACCAGGCCGTACGGGCCGCGCTCGACGAC
GGCACCCGGGTCTTCCTGGAGGTCTCCTCGCACCCGGTGGTGGCCCACTCCATCACCGAG
ACCGCACTGGACGCCGGGGTCCCCGACGCCCACGTGGCGATCACCCTGCGCAGGGAACAG
CCGGAGCAGCGCACCGTGCTGGCCAACCTGGCCCGGCTCCACAGCCTCGGCACTCCGGTC
ACCTGGTCCTACGACGGTGACCTCGTGGACGTCCCCGCGGTGCGCTGGCAGCACAAGCCG
TACTGGATCTTCCCGGACACCGCACCGGAACAGGGCGCGGGCCTGGGCCACGACCCGCAG
ACCCACACGCTGATCGGCGCGCGCACCACCGTCGCCAGCGCTCCGGTCCAGCGCGTCTGG
CAGACCGAACTGCACATGGAGAACCGTCCGTACGCGCAGTCGCACAAGGTCGTCGGCGTG
GAGACGGTGCCCGCGTCCGTGGTGCTCAACTCGTTCATCACCGCCGCCACCAACGAGGGC
GAACGGGCCTGCGGACTGCGCGACATCGTCTTCCGCATCCCGCTGGCCGCGCACCCGACA
CGGGTGGTCCAGGTGGTGCTGGAACAGGACAAGGTGCGCATCGCCAGCCGCATCAAGCGG
GACCAGGAGAGCGGCGGCGTCCGCGACGACGAGTGGCTCACCCACACCACCGCGACCGTC
GTGCACGAGCCGGAGGTCGGGGCGCGTCCGATGGAGGACCCGGACGTCATCCGGGCCCGC
TGCCCGGTGTCCTGGACCTGGGCCAAGGTCGACGGGATCTTCCGGACCATGGGCGTGGAC
GGGTACACCTTCCCCTGGGTCGTGGAGGAGCTCCTGCGCGGTGAGGACGAGCAGTTCTCC
ACGATCACGGTGGACCACACGCCGAAGCTGCACCCGTCCAGCTGGACCGCGGTCGTCGAC
GCCGCGCTGACCGCCAGTGGTGTGCTGGTGATGGACGAGAACTCGAACGTCCTGCGCACC
TGCTCGCACCTGGAGTCCCTGTCCTTCGTGGGCCCCCCGCCGCCCAGGATCCACGTACAC
ACCGTGCGTGACCCGCGCACCCCGGACACCATCAGCATGACGGTGGCCGACGAGAGCGGC
GCCGTGGTCTGCGAGGCCCGAGGGCTGCGCTACGTCAAGGTCCAGGACATCGGCTCCGGC
GCGGTCGGCCCGCGCGACCTGGTGCACGAACTCGCCTGGGAGCCGGTGGAGGTCCCCGCG
GACGCGCCGGTCCCGTCCCAGGCCCTGGTGGTGGGCGGCGCCGCCGGCGGCCCCGCCCTG
GTCGAGGCGCTCACCGCACGCGGAGTGCGGGCACGGGCGGTACCGGACGCGACCGCGATC
GGCGACGCCTCGCTCACCTGCGCCGACGTGGTCGTCGTGGCGCCCGAGGCGCTGCTGCCC
GGGGAGGCACCCGAGCAGGCGGCGCGCCGGTGCGCGCAGCTGCTCGTCGACGCCGTGCAA
CAGGTGGCGGCCGTCCCGGACGAGCGGCGGCGGCCCAGGGTGTGGGCGCTCACCCGCGAG
GTACGGGCGGGCGCGACGGAGGCCGCGCTCGCGCACGCCCCGCTGTGGGGCGCCGGACGC
ATCGTCGCCGGCGAGCGCCCCGACCTGTGGGGCGGCGTGATCGACGTCGCGGAGAACGCC
GTACCCCAGCAGGTGGCCTCGCTCATCGGAGCTCTGCCGCACACCGAGGACGTACTGTCG
CTCGACTCCGAAGGCGTCACCGCCGCCCGGCTGCGCCAGGTCGCGCGGCCCGCGGAGCGC
GAACCGGTGGACTGCCGCCCCGACGGTACGTACCTGGTCACCGGCGGTCTGGGCGCGCTC
GGCCTGGAGGCCGCGCGCCATCTGGTCGAGCAGGGCGCCAGGCGTCTGGTGCTCATCGGC
CGGCGGGGTCTTCCCTCCCGGTCGCGGTGGGACCAGGTGGACGACCCCGCCGTGGCCGCG
CAGATCGCCGAGGTCGTCGCCCTGGAGGCGGCGGGCGCCACGGTGCGTGTCCTCAGCCTG
GACATCTCCGACGCCGAGGCGACGGCCCGTGCCCTGGACCCGGGCGCTCTGGACATGCCG
CCGGTCCGGGGCATCGTGCACTGCGCCGGTGTGGTCTCCGACGCGCTGGTGGAGAAGACC
GGTGCGGCGAACCTGGACACCACCATGGGCCCCAAGGCCGACGGCGCGATGGTCCTGCAC
CGGCTCTTCCCGGCCGGCACGCTGGACTTCTTCACGATGTTCTCCTCGTGCGGCCAGCTG
GCACGGCTCACCGGCCAGGTCAGCTACGCCTCCGCGAACTCCTTCCTGGACGCGCTCGCC
GCCTTGCGGCGTTCGCGGGGGGAGACGGGTACGACGAGCTTCGCGTGGGCGCAGTGGATC
GGACGCGGCATGGGCGAGACCACCGGCAGGGCGACGATCCTGGAGGCGGAGTCCCGCGGA
CTCGGTGGCATCACCGTCTCCGAGGCGCTGCGCAGCTGGGCCTACGCGGACCGGTTCGCG
CTGCCGTACGCGGCGGTGATGCGGGTCATGCCGGATCACACGCTGCCGGTGTTCTCCCAC
CTGTCCGTCACCGACGCGGGGGCGCAGAGCGCCGATGCCGGGGGCGTCGACTGGGCCACC
GTGCCGGCCGGGGAACTGCCGGAGCTGGTGCTGAAGGTGACCCACGAGCAGGTGGCCGCC
GAGCTCAACCTCGCGGTGGACGACATCGCGATCGACCAGCCCCTGCTGGAGCTCGGTGTC
GACTCCGTGCTCACCGTGGCACTCCGGGTACGGCTGCACCGGTGTTTCGCGGTGGACCTG
CCGCCGACGATCCTGTGGAGCAACCCGACGGTCCGCGCCCTCGCGGAGTTCCTCGCCGCC
GAGGTCGGCGGTGCCACCGCGGACGCCGAGGAGACCGACCCGGTCGCCGGCCTTCCCGCA
CCTCAACAGGGATCGGGAACTGCCGAGCAGCTCGACGCCGTCGCGGCCGCGGCCGGGTGA
[1] KS11..383
[1] AT546..858
[1] acetyl-CoA malonyl-CoA731..735
[1] TH865..1017
[1] KR1389..1582
[1] ACP1666..1739
[1] KS31..1149
[1] AT1636..2574
[1] acetyl-CoA malonyl-CoA2191..2205
[1] TH2593..3051
[1] KR4165..4746
[1] ACP4996..5217

close this sectionFeature

BLASTP
Database:UniProtKB:2011_09 		show BLAST table
InterPro
Database:interpro:38.0
IPR001227 Acyl transferase domain (Domain)
[545-664] 9.40000000000003e-60 G3DSA:3.40.366.10 [731-848] 9.40000000000003e-60 G3DSA:3.40.366.10
G3DSA:3.40.366.10   Ac_transferase_reg
IPR006162 Phosphopantetheine attachment site (PTM)
[1697-1712] PS00012
PS00012   PHOSPHOPANTETHEINE
IPR009081 Acyl carrier protein-like (Domain)
[1678-1738] 2.9e-12 PF00550
PF00550   PP-binding
[1666-1739] PS50075
PS50075   ACP_DOMAIN
[1663-1776] 5.20000089003336e-18 SSF47336
SSF47336   ACP_like
[1675-1743] 1.1e-14 G3DSA:1.10.1200.10
G3DSA:1.10.1200.10   ACP_like
IPR013968 Polyketide synthase, KR (Domain)
[1389-1580] 1.29999999999998e-54 PF08659
PF08659   KR
IPR014030 Beta-ketoacyl synthase, N-terminal (Domain)
[11-259] 3.39999999999996e-77 PF00109
PF00109   ketoacyl-synt
IPR014031 Beta-ketoacyl synthase, C-terminal (Domain)
[269-383] 1.09999999999999e-38 PF02801
PF02801   Ketoacyl-synt_C
IPR014043 Acyl transferase (Domain)
[546-858] 1.10000000000001e-88 PF00698
PF00698   Acyl_transf_1
IPR016035 Acyl transferase/acyl hydrolase/lysophospholipase (Domain)
[543-851] 8.69997440322062e-55 SSF52151
SSF52151   Acyl_Trfase/lysoPlipase
IPR016036 Malonyl-CoA ACP transacylase, ACP-binding (Domain)
[666-730] 7.49999605851445e-13 SSF55048
SSF55048   Malonyl_transacylase_ACP-bd
IPR016038 Thiolase-like, subgroup (Domain)
[11-269] 5.90000000000008e-83 G3DSA:3.40.47.10 [273-436] 6.59999999999996e-51 G3DSA:3.40.47.10
G3DSA:3.40.47.10   Thiolase-like_subgr
IPR016039 Thiolase-like (Domain)
[1-436] 1e-84 SSF53901
SSF53901   Thiolase-like
IPR016040 NAD(P)-binding domain (Domain)
[1389-1587] 1.3e-29 G3DSA:3.40.50.720
G3DSA:3.40.50.720   NAD(P)-bd
IPR020801 Polyketide synthase, acyl transferase domain (Domain)
[547-840] 2.2999842048857e-84 SM00827
SM00827   PKS_AT
IPR020806 Polyketide synthase, phosphopantetheine-binding domain (Domain)
[1671-1742] 2.70000183580794e-19 SM00823
SM00823   PKS_PP
IPR020842 Polyketide synthase/Fatty acid synthase, KR (Domain)
[1389-1582] 7.90003079443025e-46 SM00822
SM00822   PKS_KR
SignalP
[1-23] 0.237 Signal
Bacteria, Gram-negative   
TMHMM No significant hit
Azino_00250 Azino_00250   Azino_00270 Azino_00270
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