Reviewed, UniProtKB/Swiss-Prot P00956 (SYI_ECOLI)

Names and origin

Protein names

Recommended name:
    Isoleucyl-tRNA synthetase
    EC=6.1.1.5
Alternative name(s):
    Isoleucine--tRNA ligase
      Short name(s)=IleRS

Gene names

Name:	ileS
Synonyms:	ilvS
Ordered Locus Names:	b0026, JW0024

Organism

Escherichia coli (strain K12) [Complete proteome] [HAMAP]

Taxonomic identifier

83333 [NCBI]

Taxonomic lineage

Bacteria › Proteobacteria › Gammaproteobacteria › Enterobacteriales › Enterobacteriaceae › Escherichia

Protein attributes

Sequence length	938 AA.
Sequence status	Complete.
Sequence processing	The displayed sequence is further processed into a mature form.
Protein existence	Evidence at protein level.

General annotation (Comments)

Function	Catalyzes the attachment of isoleucine to tRNA(Ile). As IleRS can inadvertently accommodate and process structurally similar amino acids such as valine, to avoid such errors it has two additional distinct tRNA(Ile)-dependent editing activities. One activity is designated as 'pretransfer' editing and involves the hydrolysis of activated Val-AMP. The other activity is designated 'posttransfer' editing and involves deacylation of mischarged Val-tRNA(Ile).
Catalytic activity	ATP + L-isoleucine + tRNA(Ile) = AMP + diphosphate + L-isoleucyl-tRNA(Ile).
Cofactor	Binds 1 zinc ion per subunit.
Subunit structure	Monomer.
Subcellular location	Cytoplasm.
Domain	IleRS has two distinct active sites: one for aminoacylation and one for editing. The misactivated valine is translocated from the active site to the editing site, which sterically excludes the correctly activated isoleucine. The single editing site contains two valyl binding pockets, one specific for each substrate (Val-AMP or Val-tRNA(Ile)).
Miscellaneous	Strain PS102 is resistant to the antibiotic mupirocin (pseudomonic acid A), an Ile-analog that competitively inhibits activation by Ile-tRNA synthetase, thus inhibiting protein biosynthesis.
Sequence similarities	Belongs to the class-I aminoacyl-tRNA synthetase family. IleS type 1 subfamily.
Biophysicochemical properties	Kinetic parameters: KM=5 µM for isoleucine KM=0.4 mM for ATP

Ontologies

Keywords
Biological process	Antibiotic resistance Protein biosynthesis
Cellular component	Cytoplasm
Ligand	ATP-binding Metal-binding Nucleotide-binding Zinc
Molecular function	Aminoacyl-tRNA synthetase Ligase
Technical term	Complete proteome Direct protein sequencing
Gene Ontology (GO)
Molecular function	protein binding Inferred from physical interaction. Source: IntAct
Complete GO annotation...

Binary interactions

With	Entry	#Exp.	IntAct	Notes
tufA	P0A6N1	1	EBI-552928,EBI-301077

Sequence annotation (Features)

	Feature key	Position(s)	Length	Description	Graphical view
Molecule processing
	Initiator methionine	1	1	Removed
	Chain	2 – 938	937	Isoleucyl-tRNA synthetase
Regions
	Motif	58 – 68	11	"HIGH" region
	Motif	602 – 606	5	"KMSKS" region
Sites
	Metal binding	901	1	Zinc By similarity
	Metal binding	904	1	Zinc By similarity
	Metal binding	921	1	Zinc By similarity
	Metal binding	924	1	Zinc By similarity
	Binding site	561	1	Aminoacyl-adenylate By similarity
	Binding site	605	1	ATP By similarity
Natural variations
	Natural variant	594	1	F → L in strain: PS102.
Experimental info
	Mutagenesis	94	1	G → R: 6000-fold increase in Km for isoleucine and 4-fold increase in Km for ATP, with no effect on activity
	Mutagenesis	97	1	C → S: No effect on activity
	Mutagenesis	102	1	I → N: No significant effect on activity
	Mutagenesis	183	1	K → A: Abolishes translocation from the aminoacylation site to the editing site, without effect on aminoacylation activity and posttransfer editing; when associated with A-421
	Mutagenesis	241	1	T → A: Nearly the same editing activity as the wild-type
	Mutagenesis	242	1	T → A: Abolishes editing activity against valine, with little change in aminoacylation activity; when associated with A-250
	Mutagenesis	242	1	T → P: Abolishes editing activity against valine, with little change in aminoacylation activity
	Mutagenesis	243	1	T → A: Abolishes editing activity against valine, with little change in aminoacylation activity
	Mutagenesis	243	1	T → R: Abolishes pretransfer editing
	Mutagenesis	246	1	T → A: Nearly the same editing activity as the wild-type
	Mutagenesis	250	1	N → A: Abolishes editing activity against valine, with little change in aminoacylation activity
	Mutagenesis	333	1	H → A: Alters the specificity for hydrolysis of the aminoacyl tRNA ester, with no effect on pretransfer editing
	Mutagenesis	342	1	D → A or N: Strong decrease in total editing and deacylation activities. Severely deficient in translocation from the aminoacylation site to the editing site
	Mutagenesis	342	1	D → E: Reduces 2- to 3-fold the total editing activity and 2-fold the deacylation activity. Moderately reduces translocation from the aminoacylation site to the editing site
	Mutagenesis	421	1	W → A: Abolishes translocation from the aminoacylation site to the editing site, without effect on aminoacylation activity and posttransfer editing; when associated with A-183
	Sequence conflict	243 – 264	22	TPWTL…DYALV → RRGLCLPTAQSLLHQISTMR WW AA sequence Ref.1
	Sequence conflict	300 – 301	2	EL → DV AA sequence Ref.1
	Sequence conflict	587	1	R → C AA sequence Ref.1
	Sequence conflict	637	1	E → Q AA sequence Ref.1
	Sequence conflict	724	1	G → V AA sequence Ref.1
	Sequence conflict	738	1	A → P AA sequence Ref.1
	Sequence conflict	740 – 743	4	ADSV → RTVW AA sequence Ref.1
	Sequence conflict	787	1	F → L AA sequence Ref.1
	Sequence conflict	830	1	K → N AA sequence Ref.1
	Sequence conflict	867 – 869	3	GAT → DRRY AA sequence Ref.1

Sequences

	Sequence		Length	Mass (Da)
	P00956-1 [UniParc]. Last modified January 23, 2007. Version 5. Checksum: 238CEDAF461F01D5	FASTA	938	104,297
	10 20 30 40 50 60 MSDYKSTLNL PETGFPMRGD LAKREPGMLA RWTDDDLYGI IRAAKKGKKT FILHDGPPYA 70 80 90 100 110 120 NGSIHIGHSV NKILKDIIVK SKGLSGYDSP YVPGWDCHGL PIELKVEQEY GKPGEKFTAA 130 140 150 160 170 180 EFRAKCREYA ATQVDGQRKD FIRLGVLGDW SHPYLTMDFK TEANIIRALG KIIGNGHLHK 190 200 210 220 230 240 GAKPVHWCVD CRSALAEAEV EYYDKTSPSI DVAFQAVDQD ALKAKFAVSN VNGPISLVIW 250 260 270 280 290 300 TTTPWTLPAN RAISIAPDFD YALVQIDGQA VILAKDLVES VMQRIGVTDY TILGTVKGAE 310 320 330 340 350 360 LELLRFTHPF MGFDVPAILG DHVTLDAGTG AVHTAPGHGP DDYVIGQKYG LETANPVGPD 370 380 390 400 410 420 GTYLPGTYPT LDGVNVFKAN DIVVALLQEK GALLHVEKMQ HSYPCCWRHK TPIIFRATPQ 430 440 450 460 470 480 WFVSMDQKGL RAQSLKEIKG VQWIPDWGQA RIESMVANRP DWCISRQRTW GVPMSLFVHK 490 500 510 520 530 540 DTEELHPRTL ELMEEVAKRV EVDGIQAWWD LDAKEILGDE ADQYVKVPDT LDVWFDSGST 550 560 570 580 590 600 HSSVVDVRPE FAGHAADMYL EGSDQHRGWF MSSLMISTAM KGKAPYRQVL THGFTVDGQG 610 620 630 640 650 660 RKMSKSIGNT VSPQDVMNKL GADILRLWVA STDYTGEMAV SDEILKRAAD SYRRIRNTAR 670 680 690 700 710 720 FLLANLNGFD PAKDMVKPEE MVVLDRWAVG CAKAAQEDIL KAYEAYDFHE VVQRLMRFCS 730 740 750 760 770 780 VEMGSFYLDI IKDRQYTAKA DSVARRSCQT ALYHIAEALV RWMAPILSFT ADEVWGYLPG 790 800 810 820 830 840 EREKYVFTGE WYEGLFGLAD SEAMNDAFWD ELLKVRGEVN KVIEQARADK KVGGSLEAAV 850 860 870 880 890 900 TLYAEPELSA KLTALGDELR FVLLTSGATV ADYNDAPADA QQSEVLKGLK VALSKAEGEK 910 920 930 CPRCWHYTQD VGKVAEHAEI CGRCVSNVAG DGEKRKFA

References

	« Hide 'large scale' referencesShow 'large scale' references »
[1]	"Specific sequence homology and three-dimensional structure of an aminoacyl transfer RNA synthetase." Webster T., Tsai H., Kula M., Mackie G.A., Schimmel P. Science 226:1315-1317(1984) [PubMed: 6390679] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], PARTIAL PROTEIN SEQUENCE.
[2]	"Relationship of protein structure of isoleucyl-tRNA synthetase with pseudomonic acid resistance of Escherichia coli. A proposed mode of action of pseudomonic acid as an inhibitor of isoleucyl-tRNA synthetase." Yanagisawa T., Lee J.T., Wu H.C., Kawakami M. J. Biol. Chem. 269:24304-24309(1994) [PubMed: 7929087] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], PROTEIN SEQUENCE OF 2-11 AND 606-615. Strain: K12 / MC4100 / ATCC 35695 / DSM 6574 and PS102.
[3]	"Systematic sequencing of the Escherichia coli genome: analysis of the 0-2.4 min region." Yura T., Mori H., Nagai H., Nagata T., Ishihama A., Fujita N., Isono K., Mizobuchi K., Nakata A. Nucleic Acids Res. 20:3305-3308(1992) [PubMed: 1630901] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: K12.
[4]	"The complete genome sequence of Escherichia coli K-12." Blattner F.R., Plunkett G. III, Bloch C.A., Perna N.T., Burland V., Riley M., Collado-Vides J., Glasner J.D., Rode C.K., Mayhew G.F., Gregor J., Davis N.W., Kirkpatrick H.A., Goeden M.A., Rose D.J., Mau B., Shao Y. Science 277:1453-1474(1997) [PubMed: 9278503] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: K12 / MG1655 / ATCC 47076.
[5]	"Highly accurate genome sequences of Escherichia coli K-12 strains MG1655 and W3110." Hayashi K., Morooka N., Yamamoto Y., Fujita K., Isono K., Choi S., Ohtsubo E., Baba T., Wanner B.L., Mori H., Horiuchi T. Mol. Syst. Biol. 2:E1-E5(2006) [PubMed: 16738553] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA], SEQUENCE REVISION. Strain: K12 / W3110 / ATCC 27325 / DSM 5911.
[6]	"Characterization of the ileS-lsp operon in Escherichia coli. Identification of an open reading frame upstream of the ileS gene and potential promoter(s) for the ileS-lsp operon." Kamio Y., Lin C.-K., Regue M., Wu H.C. J. Biol. Chem. 260:5616-5620(1985) [PubMed: 2985604] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA] OF 1-71.
[7]	"Nucleotide sequence of the lspA gene, the structural gene for lipoprotein signal peptidase of Escherichia coli." Yu F., Yamada H., Daishima K., Mizushima S. FEBS Lett. 173:264-268(1984) [PubMed: 6378662] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA] OF 794-938.
[8]	"Nucleotide sequence of the Escherichia coli prolipoprotein signal peptidase (lsp) gene." Innis M.A., Tokunaga M., Williams M.E., Loranger J.M., Chang S.-Y., Chang S., Wu H.C. Proc. Natl. Acad. Sci. U.S.A. 81:3708-3712(1984) [PubMed: 6374664] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA] OF 795-938.
[9]	"Evidence from cassette mutagenesis for a structure-function motif in a protein of unknown structure." Clarke N.D., Lien D.C., Schimmel P. Science 240:521-523(1988) [PubMed: 3282306] [Abstract] Cited for: MUTAGENESIS OF GLY-94; CYS-97 AND ILE-102, KINETIC PARAMETERS.
[10]	"Enzyme structure with two catalytic sites for double-sieve selection of substrate." Nureki O., Vassylyev D.G., Tateno M., Shimada A., Nakama T., Fukai S., Konno M., Hendrickson T.L., Schimmel P., Yokoyama S. Science 280:578-582(1998) [PubMed: 9554847] [Abstract] Cited for: MUTAGENESIS OF THR-241; THR-243; THR-246 AND ASN-250.
[11]	"Transfer RNA-dependent translocation of misactivated amino acids to prevent errors in protein synthesis." Nomanbhoy T.K., Hendrickson T.L., Schimmel P. Mol. Cell 4:519-528(1999) [PubMed: 10549284] [Abstract] Cited for: EDITING ACTIVITY.
[12]	"Errors from selective disruption of the editing center in a tRNA synthetase." Hendrickson T.L., Nomanbhoy T.K., Schimmel P. Biochemistry 39:8180-8186(2000) [PubMed: 10889024] [Abstract] Cited for: MUTAGENESIS OF THR-242 AND ASN-250.
[13]	"Mutational separation of two pathways for editing by a class I tRNA synthetase." Hendrickson T.L., Nomanbhoy T.K., de Crecy-Lagard V., Fukai S., Nureki O., Yokoyama S., Schimmel P. Mol. Cell 9:353-362(2002) [PubMed: 11864608] [Abstract] Cited for: MUTAGENESIS OF THR-243 AND HIS-333, PRESENCE OF TWO EDITING SUBSITES.
[14]	"Blocking site-to-site translocation of a misactivatd amino acid by mutation of a class I tRNA synthetase." Bishop A.C., Nomanbhoy T.K., Schimmel P. Proc. Natl. Acad. Sci. U.S.A. 99:585-590(2002) [PubMed: 11782529] [Abstract] Cited for: MUTAGENESIS OF ASP-342, TRANSLOCATION OF MISACTIVATED VALINE.
[15]	"Interstice mutations that block site-to-site translocation of a misactivated amino acid bound to a class I tRNA synthetase." Bishop A.C., Beebe K., Schimmel P.R. Proc. Natl. Acad. Sci. U.S.A. 100:490-494(2003) [PubMed: 12515858] [Abstract] Cited for: MUTAGENESIS OF LYS-183 AND TRP-421.

Cross-references

Sequence databases
EMBLGenBankDDBJ	U00096 Genomic DNA. Translation: AAC73137.1. AP009048 Genomic DNA. Translation: BAB96595.2. M10428 Genomic DNA. Translation: AAA24606.1. X00776 Genomic DNA. Translation: CAA25352.1. K01990 Genomic DNA. Translation: AAA24091.1.
PIR	SYECIT. B64723.
RefSeq	AP_000690.1. NP_414567.1.
3D structure databases
HSSP	HSSP built from PDB template 1FFY based on UniProtKB P41972.
ModBase	Search...
Protein-protein interaction databases
DIP	DIP:10017N.
IntAct	P00956.
2-D gel databases
ECO2DBASE	F107.0. 6TH EDITION.
Genome annotation databases
GeneID	944761.
GenomeReviews	Gene locus b0026 in contig U00096_GR. Gene locus JW0024 in contig AP009048_GR.
KEGG	ecj:JW0024. eco:b0026.
Organism-specific databases
EchoBASE	EB0487.
EcoGene	EG10492. ileS.
CMR	Search...
Phylogenomic databases
HOGENOM	P00956.
Enzyme and pathway databases
BioCyc	EcoCyc:ILES-MON.
Family and domain databases
HAMAP	MF_02002. [Tree]
InterPro	IPR001412. aa-tRNA-synth_I_CS. IPR002300. aa-tRNA-synth_Ia. IPR010663. DNA_glyclase/IsotRNA_synth_Znf. IPR002301. Ile-tRNA-synt_Ia. IPR015905. Ile-tRNA-synt_Ia_N. IPR014729. Rossmann-like_a/b/a_fold. IPR013155. V/L/I-tRNA-synth_anticodon-bd. [Graphical view]
Gene3D	G3DSA:3.40.50.620. Rossmann-like_a/b/a_fold. 1 hit.
PANTHER	PTHR11946:SF9. Ile-tRNA-synt_Ia. 1 hit.
Pfam	PF08264. Anticodon_1. 1 hit. PF00133. tRNA-synt_1. 1 hit. PF06827. zf-FPG_IleRS. 1 hit. [Graphical view]
PRINTS	PR00984. TRNASYNTHILE.
TIGRFAMs	TIGR00392. ileS. 1 hit.
PROSITE	PS00178. AA_TRNA_LIGASE_I. 1 hit. [Graphical view]
ProDom	P00956. [Graphical view] [Entries sharing at least one domain]
BLOCKS	Search...
Other Resources
BindingDB	P00956.
ProtoNet	Search...

Entry information

Entry name

SYI_ECOLI

Accession

Primary (citable) accession number: P00956
Secondary accession number(s): P78038, Q59429

Entry history

Integrated into UniProtKB/Swiss-Prot:	July 21, 1986
Last sequence update:	January 23, 2007
Last modified:	July 22, 2008
This is version 100 of the entry and version 5 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation project

HAMAP (High-quality Automated and Manual Annotation of microbial Proteomes)