CARA_PECCC
ID CARA_PECCC Reviewed; 503 AA.
AC Q9XB61;
DT 16-OCT-2013, integrated into UniProtKB/Swiss-Prot.
DT 01-NOV-1999, sequence version 1.
DT 03-AUG-2022, entry version 70.
DE RecName: Full=Carbapenam-3-carboxylate synthase;
DE EC=6.3.3.6;
DE AltName: Full=Carbapenam-3-carboxylate ligase;
GN Name=carA;
OS Pectobacterium carotovorum subsp. carotovorum (Erwinia carotovora subsp.
OS carotovora).
OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales;
OC Pectobacteriaceae; Pectobacterium.
OX NCBI_TaxID=555;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RC STRAIN=ATCC 39048 / GS101 / SC 12;
RX PubMed=9402024; DOI=10.1046/j.1365-2958.1997.6001974.x;
RA McGowan S.J., Sebaihia M., O'Leary S., Hardie K.R., Williams P.,
RA Stewart G.S., Bycroft B.W., Salmond G.P.;
RT "Analysis of the carbapenem gene cluster of Erwinia carotovora: definition
RT of the antibiotic biosynthetic genes and evidence for a novel beta-lactam
RT resistance mechanism.";
RL Mol. Microbiol. 26:545-556(1997).
RN [2]
RP FUNCTION, CATALYTIC ACTIVITY, AND BIOPHYSICOCHEMICAL PROPERTIES.
RX PubMed=12820893; DOI=10.1021/bi034361d;
RA Gerratana B., Stapon A., Townsend C.A.;
RT "Inhibition and alternate substrate studies on the mechanism of carbapenam
RT synthetase from Erwinia carotovora.";
RL Biochemistry 42:7836-7847(2003).
RN [3]
RP FUNCTION, CATALYTIC ACTIVITY, BIOPHYSICOCHEMICAL PROPERTIES, AND
RP MUTAGENESIS OF LYS-443.
RX PubMed=17658887; DOI=10.1021/bi0618464;
RA Arnett S.O., Gerratana B., Townsend C.A.;
RT "Rate-limiting steps and role of active site Lys443 in the mechanism of
RT carbapenam synthetase.";
RL Biochemistry 46:9337-9345(2007).
RN [4]
RP FUNCTION, CATALYTIC ACTIVITY, ACTIVE SITE, AND MUTAGENESIS OF TYR-345 AND
RP GLU-380.
RX PubMed=19371088; DOI=10.1021/bi900432n;
RA Raber M.L., Arnett S.O., Townsend C.A.;
RT "A conserved tyrosyl-glutamyl catalytic dyad in evolutionarily linked
RT enzymes: carbapenam synthetase and beta-lactam synthetase.";
RL Biochemistry 48:4959-4971(2009).
RN [5]
RP X-RAY CRYSTALLOGRAPHY (2.30 ANGSTROMS) IN COMPLEX WITH SUBSTRATE AND ATP
RP ANALOG, AND SUBUNIT.
RX PubMed=12890666; DOI=10.1074/jbc.m307901200;
RA Miller M.T., Gerratana B., Stapon A., Townsend C.A., Rosenzweig A.C.;
RT "Crystal structure of carbapenam synthetase (CarA).";
RL J. Biol. Chem. 278:40996-41002(2003).
CC -!- FUNCTION: Involved in the biosynthesis of carbapenam-3-carboxylate, a
CC beta-lactam antibiotic of the carbapenem class. Catalyzes the ATP-
CC dependent formation of (3S,5S)-carbapenam-3-carboxylate from (2S,5S)-5-
CC carboxymethylproline. {ECO:0000269|PubMed:12820893,
CC ECO:0000269|PubMed:17658887, ECO:0000269|PubMed:19371088}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=(2S,5S)-5-carboxymethylproline + ATP = (3S,5S)-carbapenam-3-
CC caboxylate + AMP + diphosphate + H(+); Xref=Rhea:RHEA:36703,
CC ChEBI:CHEBI:15378, ChEBI:CHEBI:30616, ChEBI:CHEBI:33019,
CC ChEBI:CHEBI:73939, ChEBI:CHEBI:73962, ChEBI:CHEBI:456215; EC=6.3.3.6;
CC Evidence={ECO:0000269|PubMed:12820893, ECO:0000269|PubMed:17658887,
CC ECO:0000269|PubMed:19371088};
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=0.11 mM for ATP {ECO:0000269|PubMed:12820893,
CC ECO:0000269|PubMed:17658887};
CC KM=0.23 mM for (2S,5S)-5-carboxymethylproline
CC {ECO:0000269|PubMed:12820893, ECO:0000269|PubMed:17658887};
CC Note=kcat is 0.28 sec(-1).;
CC pH dependence:
CC Optimum pH is 8.5. {ECO:0000269|PubMed:12820893,
CC ECO:0000269|PubMed:17658887};
CC -!- PATHWAY: Antibiotic biosynthesis; carbapenem biosynthesis.
CC -!- SUBUNIT: Homotetramer. {ECO:0000269|PubMed:12890666}.
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DR EMBL; U17224; AAD38229.1; -; Genomic_DNA.
DR PDB; 1Q15; X-ray; 2.30 A; A/B/C/D=1-503.
DR PDB; 1Q19; X-ray; 2.40 A; A/B/C/D=1-503.
DR PDBsum; 1Q15; -.
DR PDBsum; 1Q19; -.
DR AlphaFoldDB; Q9XB61; -.
DR SMR; Q9XB61; -.
DR DrugBank; DB03215; (2s,5s)-5-Carboxymethylproline.
DR DrugBank; DB02596; alpha,beta-Methyleneadenosine 5'-triphosphate.
DR KEGG; ag:AAD38229; -.
DR BioCyc; MetaCyc:MON-13572; -.
DR BRENDA; 6.3.3.6; 2140.
DR UniPathway; UPA00182; -.
DR EvolutionaryTrace; Q9XB61; -.
DR GO; GO:0004066; F:asparagine synthase (glutamine-hydrolyzing) activity; IEA:InterPro.
DR GO; GO:0005524; F:ATP binding; IEA:UniProtKB-KW.
DR GO; GO:0017000; P:antibiotic biosynthetic process; IEA:UniProtKB-KW.
DR GO; GO:0006529; P:asparagine biosynthetic process; IEA:InterPro.
DR CDD; cd01991; Asn_Synthase_B_C; 1.
DR Gene3D; 3.40.50.620; -; 1.
DR Gene3D; 3.60.20.10; -; 1.
DR InterPro; IPR001962; Asn_synthase.
DR InterPro; IPR015230; CarA_N.
DR InterPro; IPR029055; Ntn_hydrolases_N.
DR InterPro; IPR014729; Rossmann-like_a/b/a_fold.
DR Pfam; PF00733; Asn_synthase; 2.
DR Pfam; PF09147; DUF1933; 1.
DR SUPFAM; SSF56235; SSF56235; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Antibiotic biosynthesis; ATP-binding; Ligase;
KW Nucleotide-binding.
FT CHAIN 1..503
FT /note="Carbapenam-3-carboxylate synthase"
FT /id="PRO_0000424221"
FT DOMAIN 228..443
FT /note="Asparagine synthetase"
FT ACT_SITE 345
FT /evidence="ECO:0000305|PubMed:19371088"
FT ACT_SITE 380
FT /note="Proton acceptor"
FT /evidence="ECO:0000305|PubMed:19371088"
FT BINDING 244..251
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 270
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 344..348
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 346
FT /ligand="substrate"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 371
FT /ligand="substrate"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 374
FT /ligand="substrate"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 421
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:12890666"
FT BINDING 444..446
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:12890666"
FT MUTAGEN 345
FT /note="Y->A: Loss of activity."
FT /evidence="ECO:0000269|PubMed:19371088"
FT MUTAGEN 345
FT /note="Y->F: Reduces catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19371088"
FT MUTAGEN 380
FT /note="E->A: Loss of activity."
FT /evidence="ECO:0000269|PubMed:19371088"
FT MUTAGEN 380
FT /note="E->D: Reduces catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19371088"
FT MUTAGEN 380
FT /note="E->Q: Reduces catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19371088"
FT MUTAGEN 443
FT /note="K->A,M: Loss of activity."
FT /evidence="ECO:0000269|PubMed:17658887"
FT STRAND 5..9
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 13..20
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 25..30
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 33..38
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 43..47
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 49..57
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 62..69
FT /evidence="ECO:0007829|PDB:1Q15"
FT TURN 70..72
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 74..78
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 81..92
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 94..99
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 102..109
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 115..119
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 121..125
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 128..139
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 141..148
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 158..161
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 177..179
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 182..190
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 196..203
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 216..234
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 235..237
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 239..244
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 249..258
FT /evidence="ECO:0007829|PDB:1Q15"
FT TURN 259..261
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 263..272
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 278..288
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 291..297
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 299..313
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 318..334
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 338..342
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 347..350
FT /evidence="ECO:0007829|PDB:1Q15"
FT TURN 351..354
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 364..378
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 384..388
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 392..394
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 400..407
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 411..413
FT /evidence="ECO:0007829|PDB:1Q15"
FT STRAND 419..421
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 422..430
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 436..439
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 446..449
FT /evidence="ECO:0007829|PDB:1Q19"
FT HELIX 452..460
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 467..482
FT /evidence="ECO:0007829|PDB:1Q15"
FT HELIX 488..490
FT /evidence="ECO:0007829|PDB:1Q19"
FT HELIX 493..500
FT /evidence="ECO:0007829|PDB:1Q15"
SQ SEQUENCE 503 AA; 55998 MW; 3BB1DBBFF352DBCC CRC64;
MSNSFCVVYK GSDTDINNIQ RDFDGKGEAL SNGYLFIEQN GHYQKCEMER GTAYLIGSLY
NRTFLIGLAG VWEGEAYLAN DAELLALLFT RLGANALALA EGDFCFFIDE PNGELTVITE
SRGFSPVHVV QGKKAWMTNS LKLVTAAEGE GALWFEEEAL VCQSLMRADT YTPVKNAQRL
KPGAVHVLTH DSEGYSFVES RTLTTPASNQ LLALPREPLL ALIDRYLNAP LEDLAPRFDT
VGIPLSGGLD SSLVTALASR HFKKLNTYSI GTELSNEFEF SQQVADALGT HHQMKILSET
EVINGIIESI YYNEIFDGLS AEIQSGLFNV YRQAQGQVSC MLTGYGSDLL FGGILKPGAQ
YDNPNQLLAE QVYRTRWTGE FATHGASCYG IDIRHPFWSH SLISLCHALH PDYKIFDNEV
KNILREYADS LQLLPKDIVW RKKIGIHEGS SVNQAFANVL GSTVDNYQTK SRFTYRVYQA
FLRGRLSITD VTPSQLKDLI KKD
