Rapam_00200 : CDS information

Location

Organism	Streptomyces rapamycinicus
Strain	NRRL 5491
Entry name	Rapamycin
Contig	X86780
Start / Stop / Direction	95,430 / 96,434 / + [in whole cluster] 95,430 / 96,434 / + [in contig]
Location	95430..96434 [in whole cluster] 95430..96434 [in contig]
Type	CDS
Length	1,005 bp (334 aa)

Annotation

Category	2.2 modification addition of starter unit
Product	putative chorismatase putative chorismate hydrolase
Product (GenBank)	pteridine-dependent dioxygenase
Gene
Gene (GenBank)	rapK
EC number
Keyword	4,5-dihydroxycyclohex-1-ene carboxylic acid
Note
Note (GenBank)
Reference	ACC Q54305 PmId [7644502] The biosynthetic gene cluster for the polyketide immunosuppressant rapamycin. (Proc Natl Acad Sci U S A. , 1995) [8635730] Organisation of the biosynthetic gene cluster for rapamycin in Streptomyces hygroscopicus: analysis of genes flanking the polyketide synthase. (Gene. , 1996) [15127450] Isolation and characterization of pre-rapamycin, the first macrocyclic intermediate in the biosynthesis of the immunosuppressant rapamycin by S. hygroscopicus. (Angew Chem Int Ed Engl. , 2004) [21383123] Biosynthesis of the immunosuppressants FK506, FK520, and rapamycin involves a previously undescribed family of enzymes acting on chorismate. (Proc Natl Acad Sci U S A. , 2011) comment [PMID: 7644502](1995) [PMID: 8635730](1996) cluster genesの報告。 -- [PMID:21383123](2011) rapamycin生合成において、(4R,5R)-4,5-dihydroxycyclohex-1-enecarboxylic acid(=DHCHC)がstarter unit前駆体であり、その産生にはrapKが必須なことがわかっている(ref[PMID: 15127450])。 rapK deletion mutantはrapamycin非産生。 rapK, FK506 or FK520 gene cluster由来fkbOの発現や、DHCHC添加で補完され、rapamycin産生を回復する。よってrapK, fkbOは同じ機能を持つ。 FK520 gene clusterのfkbOをE.coli発現、精製。このタンパクでChorismateは(4R,5R)-4,5-dihydroxycyclohexa-1,5-dienecarboxylic acid(=DCDC)へ変換されたことを、産物解析により確認。 kinetic parametersは反応産物pyruvateをモニターして測定。 chorismate + H2O → DCDC + pyruvate rapK deletion mutantはstarter unitが3-hydroxybenzoateのBC325を蓄積。 uncharacterized hyg clusterにあるhyg5がchorismate→3-hydroxybenzoateへの変換を担うことも確認している。
Related Reference	ACC Q9KID9 NITE Asco_00150 PmId [21383123] Biosynthesis of the immunosuppressants FK506, FK520, and rapamycin involves a previously undescribed family of enzymes acting on chorismate. (Proc Natl Acad Sci U S A. , 2011) [24036425] Mechanistic implications for the chorismatase FkbO based on the crystal structure. (J Mol Biol. , 2014) [26247872] Chorismatase Mechanisms Reveal Fundamentally Different Types of Reaction in a Single Conserved Protein Fold. (J Am Chem Soc. , 2015) comment BLAST id73%, 1e-135 Streptomyces hygroscopicus subsp. ascomyceticus_fkbO [Asco_00150]chorismatase(chorismate hydrolase) -- [PMID: 21383123](2011) FK520 gene clusterのfkbOをE.coli発現、精製。このタンパクでChorismateは(4R,5R)-4,5-dihydroxycyclohexa-1,5-dienecarboxylic acid(=DCDC)へ変換されたことを、産物解析により確認。 kinetic parametersは反応産物pyruvateをモニターして測定。 chorismate + H2O → DCDC + pyruvate rapamycin生合成において、(4R,5R)-4,5-dihydroxycyclohex-1-enecarboxylic acid(=DHCHC)がstarter unit前駆体であり、その産生にはrapKが必須なことがわかっている。 rapK deletion mutantはrapamycin非産生。 FK506 or FK520 gene cluster由来fkbOで補完され、rapamycin産生回復。 --- [PMID:24036425](2013) 結晶構造解析から、FkbOの反応モデルを提唱。 active-site変異体の活性測定でそれを確認している。

Sequence

>putative chorismatase [pteridine-dependent dioxygenase]
MRQLTPPVTAPYCRFEKLGASDLDGDETLLGVIEHRTGHTGVSLAEGCPRTAVHTTTRED
ESFAEAWHAEGPKESSRHDGVAWARTPDYLFGVARVPEGGRYAAGTAAVYTGIFDLIGTL
GYPSLARTWNYVSGINTPNADGLEVYRDFCVGRAEALDARGIDPATMPAATGIGAHGARI
TCYFIAARAGDRVNMENPAVLTAHRYPQRYGPRPPVFSGHLALAAGGGRLFVSATAGIVG
QETVHHGDVAAQCEVSLENIARVIGAENLGRHGLRRGYALADVDHLKVYVRHREDISTVR
RICAERLSREATVAVLHTDIARTDLLVEIEGVVA

>putative chorismatase [pteridine-dependent dioxygenase]
GTGAGGCAATTGACTCCGCCGGTCACGGCACCGTACTGCCGCTTCGAGAAGCTCGGCGCC
TCCGATCTCGACGGCGACGAGACACTTCTCGGCGTCATCGAACATCGCACCGGACACACC
GGGGTTTCACTGGCCGAGGGCTGTCCCCGGACGGCCGTACACACGACGACCCGCGAGGAC
GAGTCGTTCGCCGAGGCGTGGCACGCGGAGGGGCCGAAGGAGTCGTCGAGGCATGATGGC
GTCGCCTGGGCTCGGACACCTGACTACCTCTTCGGTGTCGCGCGGGTGCCCGAGGGCGGC
CGGTACGCGGCCGGCACCGCGGCCGTCTACACCGGAATCTTCGACCTGATCGGGACGCTG
GGGTACCCCAGTCTGGCCCGCACCTGGAACTACGTCAGCGGAATCAACACGCCGAACGCC
GATGGCCTCGAGGTCTACCGGGACTTCTGTGTGGGCCGCGCCGAGGCGCTGGACGCCCGT
GGGATCGACCCGGCGACCATGCCGGCGGCGACCGGCATCGGCGCCCACGGCGCGCGCATC
ACGTGCTACTTCATCGCCGCACGCGCCGGTGACCGGGTCAACATGGAGAACCCGGCCGTG
CTCACGGCTCACCGCTACCCGCAGCGGTACGGCCCCCGCCCGCCGGTCTTCTCCGGCCAC
CTGGCTCTCGCCGCCGGGGGCGGACGGCTCTTCGTCTCCGCGACCGCCGGCATCGTCGGT
CAGGAGACGGTGCACCACGGCGACGTCGCCGCGCAATGCGAGGTGTCGCTCGAGAACATC
GCCCGGGTGATCGGTGCCGAGAACCTGGGACGGCACGGGCTCCGCCGGGGCTACGCCCTC
GCCGACGTCGACCATTTGAAGGTCTATGTACGCCACCGCGAGGATATTTCGACCGTACGT
CGGATCTGCGCCGAGCGCCTCTCCCGCGAGGCCACGGTTGCCGTACTGCACACCGACATC
GCGCGCACCGATCTGCTGGTCGAGATCGAGGGTGTGGTGGCGTGA

Feature

BLASTP Database:UniProtKB:2011_09	show BLAST table
InterPro Database:interpro:38.0	IPR013813 Endoribonuclease L-PSP/chorismate mutase-like (Domain) G3DSA:3.30.1330.40 Endoribo_LPSP/chorism_mut-like SSF55298 YjgF/chorismate_mutase-like
SignalP	No significant hit
TMHMM	No significant hit

Rapam_00190 Rapam_00210