SUCPP_THETC
ID SUCPP_THETC Reviewed; 488 AA.
AC D9TT09;
DT 26-NOV-2014, integrated into UniProtKB/Swiss-Prot.
DT 05-OCT-2010, sequence version 1.
DT 03-AUG-2022, entry version 67.
DE RecName: Full=Sucrose 6(F)-phosphate phosphorylase {ECO:0000305|PubMed:24599311};
DE EC=2.4.1.329 {ECO:0000269|PubMed:24599311};
DE AltName: Full=Sucrose 6'-phosphate phosphorylase {ECO:0000303|PubMed:24599311};
DE Short=SPP {ECO:0000303|PubMed:24599311};
GN Name=spp {ECO:0000303|PubMed:24599311}; OrderedLocusNames=Tthe_1921;
OS Thermoanaerobacterium thermosaccharolyticum (strain ATCC 7956 / DSM 571 /
OS NCIMB 9385 / NCA 3814 / NCTC 13789 / WDCM 00135 / 2032) (Clostridium
OS thermosaccharolyticum).
OC Bacteria; Firmicutes; Clostridia; Thermoanaerobacterales;
OC Thermoanaerobacterales Family III. Incertae Sedis; Thermoanaerobacterium.
OX NCBI_TaxID=580327;
RN [1]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=ATCC 7956 / DSM 571 / NCIMB 9385 / NCA 3814 / NCTC 13789 / WDCM
RC 00135 / 2032;
RG US DOE Joint Genome Institute;
RA Lucas S., Copeland A., Lapidus A., Cheng J.-F., Bruce D., Goodwin L.,
RA Pitluck S., Teshima H., Detter J.C., Han C., Tapia R., Land M., Hauser L.,
RA Chang Y.-J., Jeffries C., Kyrpides N., Ivanova N., Mikhailova N.,
RA Hemme C.L., Woyke T.;
RT "Complete sequence of Thermoanaerobacterium thermosaccharolyticum DSM
RT 571.";
RL Submitted (AUG-2010) to the EMBL/GenBank/DDBJ databases.
RN [2]
RP FUNCTION, CATALYTIC ACTIVITY, SUBSTRATE SPECIFICITY, BIOPHYSICOCHEMICAL
RP PROPERTIES, MUTAGENESIS OF ARG-134; ARG-135; LYS-301 AND HIS-344, AND
RP 3D-STRUCTURE MODELING.
RX PubMed=24599311; DOI=10.1007/s00253-014-5621-y;
RA Verhaeghe T., Aerts D., Diricks M., Soetaert W., Desmet T.;
RT "The quest for a thermostable sucrose phosphorylase reveals sucrose 6'-
RT phosphate phosphorylase as a novel specificity.";
RL Appl. Microbiol. Biotechnol. 98:7027-7037(2014).
RN [3]
RP MUTAGENESIS OF ARG-134, BIOTECHNOLOGY, AND FUNCTION.
RX PubMed=26074151; DOI=10.1002/anie.201503605;
RA Dirks-Hofmeister M.E., Verhaeghe T., De Winter K., Desmet T.;
RT "Creating space for large acceptors: rational biocatalyst design for
RT resveratrol glycosylation in an aqueous system.";
RL Angew. Chem. Int. Ed. 54:9289-9292(2015).
RN [4] {ECO:0007744|PDB:6S9V}
RP X-RAY CRYSTALLOGRAPHY (1.83 ANGSTROMS), SUBUNIT, AND ACTIVE SITE.
RX PubMed=31405215; DOI=10.3390/ijms20163906;
RA Franceus J., Capra N., Desmet T., Thunnissen A.W.H.;
RT "Structural Comparison of a Promiscuous and a Highly Specific Sucrose 6F-
RT Phosphate Phosphorylase.";
RL Int. J. Mol. Sci. 20:0-0(2019).
CC -!- FUNCTION: Catalyzes the reversible phosphorolysis of sucrose 6(F)-
CC phosphate into alpha-D-glucose 1-phosphate (Glc1P) and D-fructose 6-
CC phosphate. May be involved in a new pathway for the degradation of
CC sucrose, which could become phosphorylated on its fructose moiety
CC during uptake via a PTS system. To a lesser extent, can also reversibly
CC act on sucrose in vitro (PubMed:24599311). Is also able to catalyze
CC transglycosylation reactions in vitro (PubMed:26074151).
CC {ECO:0000269|PubMed:24599311, ECO:0000269|PubMed:26074151}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=phosphate + sucrose 6(F)-phosphate = alpha-D-glucose 1-
CC phosphate + beta-D-fructose 6-phosphate; Xref=Rhea:RHEA:38863,
CC ChEBI:CHEBI:43474, ChEBI:CHEBI:57634, ChEBI:CHEBI:57723,
CC ChEBI:CHEBI:58601; EC=2.4.1.329;
CC Evidence={ECO:0000269|PubMed:24599311};
CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:38864;
CC Evidence={ECO:0000305|PubMed:24599311};
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=6.9 mM for phosphate (at 55 degrees Celsius and pH 6.5)
CC {ECO:0000269|PubMed:24599311};
CC KM=12.7 mM for sucrose 6(F)-phosphate (at 55 degrees Celsius and pH
CC 6.5) {ECO:0000269|PubMed:24599311};
CC KM=76.5 mM for sucrose (at 55 degrees Celsius and pH 6.5)
CC {ECO:0000269|PubMed:24599311};
CC KM=15.1 mM for D-fructose 6-phosphate (at 55 degrees Celsius and pH
CC 6.0) {ECO:0000269|PubMed:24599311};
CC KM=15.6 mM for alpha-D-glucose 1-phosphate (at 55 degrees Celsius and
CC pH 6.0) {ECO:0000269|PubMed:24599311};
CC KM=41.6 mM for D-fructose (at 55 degrees Celsius and pH 6.0)
CC {ECO:0000269|PubMed:24599311};
CC Note=kcat is 82.6 sec(-1) for the phosphorolysis of sucrose 6(F)-
CC phosphate (at 55 degrees Celsius and pH 6.5). kcat is 66.2 sec(-1)
CC for the phosphorolysis of sucrose (at 55 degrees Celsius and pH 6.5).
CC kcat is 24.2 sec(-1) for the synthetic direction with alpha-D-glucose
CC 1-phosphate and D-fructose 6-phosphate as substrates (at 55 degrees
CC Celsius and pH 6.0). kcat is 14.4 sec(-1) for the synthetic direction
CC with alpha-D-glucose 1-phosphate and D-fructose as substrates (at 55
CC degrees Celsius and pH 6.0). {ECO:0000269|PubMed:24599311};
CC pH dependence:
CC Optimum pH is 6 and 6.5 for the synthetic and phosphorolytic
CC reaction, respectively. Thermostable. Has a half-life of 60 hours at
CC 60 degrees Celsius. {ECO:0000269|PubMed:24599311};
CC Temperature dependence:
CC Optimum temperature is 55 degrees Celsius.
CC {ECO:0000269|PubMed:24599311};
CC -!- SUBUNIT: Monomer. {ECO:0000269|PubMed:31405215}.
CC -!- BIOTECHNOLOGY: The mutant Ala-134 is a promising new biocatalyst for
CC glycosylation reactions on bulky phenolic acceptors.
CC {ECO:0000305|PubMed:26074151}.
CC -!- SIMILARITY: Belongs to the glycosyl hydrolase 13 family. Sucrose
CC phosphorylase subfamily. {ECO:0000305}.
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DR EMBL; CP002171; ADL69407.1; -; Genomic_DNA.
DR RefSeq; WP_013298373.1; NC_014410.1.
DR PDB; 6S9V; X-ray; 1.83 A; A/B=1-488.
DR PDBsum; 6S9V; -.
DR AlphaFoldDB; D9TT09; -.
DR SMR; D9TT09; -.
DR STRING; 580327.Tthe_1921; -.
DR CAZy; GH13; Glycoside Hydrolase Family 13.
DR EnsemblBacteria; ADL69407; ADL69407; Tthe_1921.
DR KEGG; ttm:Tthe_1921; -.
DR eggNOG; COG0366; Bacteria.
DR HOGENOM; CLU_021358_1_0_9; -.
DR OMA; PNATQFT; -.
DR OrthoDB; 1573900at2; -.
DR BioCyc; MetaCyc:MON-18710; -.
DR BRENDA; 2.4.1.329; 1533.
DR SABIO-RK; D9TT09; -.
DR Proteomes; UP000001626; Chromosome.
DR GO; GO:0004645; F:1,4-alpha-oligoglucan phosphorylase activity; IEA:InterPro.
DR GO; GO:0016758; F:hexosyltransferase activity; IDA:UniProtKB.
DR GO; GO:0044262; P:cellular carbohydrate metabolic process; IEA:InterPro.
DR CDD; cd11355; AmyAc_Sucrose_phosphorylase; 1.
DR Gene3D; 3.90.400.10; -; 1.
DR InterPro; IPR006047; Glyco_hydro_13_cat_dom.
DR InterPro; IPR017853; Glycoside_hydrolase_SF.
DR InterPro; IPR045857; O16G_dom_2.
DR InterPro; IPR016377; Sucrose_GGa_phosphorylase-rel.
DR InterPro; IPR022527; Sucrose_phospho.
DR Pfam; PF00128; Alpha-amylase; 1.
DR PIRSF; PIRSF003059; Sucrose_phosphorylase; 1.
DR SMART; SM00642; Aamy; 1.
DR SUPFAM; SSF51445; SSF51445; 1.
DR TIGRFAMs; TIGR03852; sucrose_gtfA; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Carbohydrate metabolism; Glycosyltransferase;
KW Reference proteome; Transferase.
FT CHAIN 1..488
FT /note="Sucrose 6(F)-phosphate phosphorylase"
FT /id="PRO_0000430937"
FT ACT_SITE 197
FT /note="Nucleophile"
FT /evidence="ECO:0000305|PubMed:31405215"
FT ACT_SITE 238
FT /note="Proton donor/acceptor"
FT /evidence="ECO:0000305|PubMed:31405215"
FT BINDING 49
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT BINDING 87
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT BINDING 195..197
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT BINDING 238
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT BINDING 295..296
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT BINDING 342..345
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT BINDING 399
FT /ligand="sucrose 6(F)-phosphate"
FT /ligand_id="ChEBI:CHEBI:57723"
FT /evidence="ECO:0000250|UniProtKB:A0ZZH6"
FT SITE 296
FT /note="Transition state stabilizer"
FT /evidence="ECO:0000305|PubMed:31405215"
FT MUTAGEN 134
FT /note="R->A: Shows a decreased ratio of activity on
FT phosphorylated fructose over fructose. Displays increased
FT transglycosylation activity on a broad range of bulky
FT acceptors."
FT /evidence="ECO:0000269|PubMed:24599311,
FT ECO:0000269|PubMed:26074151"
FT MUTAGEN 135
FT /note="R->A: Displays similar activity as the wild-type."
FT /evidence="ECO:0000269|PubMed:24599311"
FT MUTAGEN 301
FT /note="K->A: Displays similar activity as the wild-type."
FT /evidence="ECO:0000269|PubMed:24599311"
FT MUTAGEN 344
FT /note="H->Y: Shows a decreased ratio of activity on
FT phosphorylated fructose over fructose."
FT /evidence="ECO:0000269|PubMed:24599311"
FT STRAND 8..11
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 20..30
FT /evidence="ECO:0007829|PDB:6S9V"
FT TURN 32..34
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 36..40
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 46..48
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 49..51
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 53..55
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 57..59
FT /evidence="ECO:0007829|PDB:6S9V"
FT TURN 62..64
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 67..74
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 77..83
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 85..89
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 93..101
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 102..104
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 108..110
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 114..116
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 125..128
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 134..143
FT /evidence="ECO:0007829|PDB:6S9V"
FT TURN 145..147
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 150..154
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 159..162
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 164..168
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 173..188
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 193..196
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 199..201
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 210..212
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 216..230
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 234..237
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 243..251
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 255..257
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 261..271
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 275..281
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 288..290
FT /evidence="ECO:0007829|PDB:6S9V"
FT TURN 301..307
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 310..322
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 326..329
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 333..335
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 343..348
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 350..353
FT /evidence="ECO:0007829|PDB:6S9V"
FT TURN 354..356
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 358..370
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 371..378
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 379..382
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 389..395
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 398..402
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 408..414
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 418..432
FT /evidence="ECO:0007829|PDB:6S9V"
FT HELIX 434..437
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 438..442
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 449..456
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 459..466
FT /evidence="ECO:0007829|PDB:6S9V"
FT TURN 467..469
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 472..477
FT /evidence="ECO:0007829|PDB:6S9V"
FT STRAND 483..487
FT /evidence="ECO:0007829|PDB:6S9V"
SQ SEQUENCE 488 AA; 56716 MW; A6F1B6D7F79B3168 CRC64;
MALKNKVQLI TYPDSLGGNL KTLNDVLEKY FSDVFGGVHI LPPFPSSGDR GFAPITYSEI
EPKFGTWYDI KKMAENFDIL LDLMVNHVSR RSIYFQDFLK KGRKSEYADM FITLDKLWKD
GKPVKGDIEK MFLRRTLPYS TFKIEETGEE EKVWTTFGKT DPSEQIDLDV NSHLVREFLL
EVFKTFSNFG VKIVRLDAVG YVIKKIGTSC FFVEPEIYEF LDWAKGQAAS YGIELLLEVH
SQFEVQYKLA ERGFLIYDFI LPFTVLYTLI NKSNEMLYHY LKNRPINQFT MLDCHDGIPV
KPDLDGLIDT KKAKEVVDIC VQRGANLSLI YGDKYKSEDG FDVHQINCTY YSALNCDDDA
YLAARAIQFF TPGIPQVYYV GLLAGVNDFE AVKKTKEGRE INRHNYGLKE IEESVQKNVV
QRLLKLIRFR NEYEAFNGEF FIEDCRKDEI RLTWKKDDKR CSLFIDLKTY KTTIDYINEN
GEEVKYLV
