SALR_PAPBR
ID SALR_PAPBR Reviewed; 311 AA.
AC A4UHT7;
DT 09-FEB-2010, integrated into UniProtKB/Swiss-Prot.
DT 15-MAY-2007, sequence version 1.
DT 03-AUG-2022, entry version 52.
DE RecName: Full=Salutaridine reductase {ECO:0000312|EMBL:ABO93462.1};
DE EC=1.1.1.248;
GN Name=SALR {ECO:0000312|EMBL:ABO93462.1};
OS Papaver bracteatum (Great scarlet poppy).
OC Eukaryota; Viridiplantae; Streptophyta; Embryophyta; Tracheophyta;
OC Spermatophyta; Magnoliopsida; Ranunculales; Papaveraceae; Papaveroideae;
OC Papaver.
OX NCBI_TaxID=215227;
RN [1] {ECO:0000305, ECO:0000312|EMBL:ABO93462.1}
RP NUCLEOTIDE SEQUENCE [MRNA], FUNCTION, CATALYTIC ACTIVITY, ACTIVITY
RP REGULATION, BIOPHYSICOCHEMICAL PROPERTIES, AND MUTAGENESIS OF ARG-44;
RP ARG-48; PHE-104; VAL-106; ASN-152; SER-180; TYR-236; LYS-240; LEU-266;
RP MET-271 AND ASN-272.
RX PubMed=17337529; DOI=10.1104/pp.106.095166;
RA Geissler R., Brandt W., Ziegler J.;
RT "Molecular modeling and site-directed mutagenesis reveal the
RT benzylisoquinoline binding site of the short-chain dehydrogenase/reductase
RT salutaridine reductase.";
RL Plant Physiol. 143:1493-1503(2007).
RN [2] {ECO:0000305}
RP FUNCTION, CATALYTIC ACTIVITY, ACTIVITY REGULATION, BIOPHYSICOCHEMICAL
RP PROPERTIES, BIOTECHNOLOGY, AND MUTAGENESIS OF PHE-104; VAL-106; ASP-107;
RP SER-181; THR-182; LEU-185; LYS-186; LEU-266; MET-271; ASN-272 AND ILE-275.
RX PubMed=19648114; DOI=10.1074/jbc.m109.030957;
RA Ziegler J., Brandt W., Geissler R., Facchini P.J.;
RT "Removal of substrate inhibition and increase in maximal velocity in the
RT short chain dehydrogenase/reductase salutaridine reductase involved in
RT morphine biosynthesis.";
RL J. Biol. Chem. 284:26758-26767(2009).
CC -!- FUNCTION: Involved in biosynthesis of morphinan-type benzylisoquinoline
CC alkaloids. Catalyzes the stereospecific conversion of salutaridine to
CC salutaridinol. {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=(7S)-salutaridinol + NADP(+) = H(+) + NADPH + salutaridine;
CC Xref=Rhea:RHEA:10108, ChEBI:CHEBI:15378, ChEBI:CHEBI:57783,
CC ChEBI:CHEBI:58061, ChEBI:CHEBI:58349, ChEBI:CHEBI:58463;
CC EC=1.1.1.248; Evidence={ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC -!- ACTIVITY REGULATION: Subject to substrate inhibition at salutaridine
CC concentrations higher than 20 to 30 uM. {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114}.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=7.9 uM for salutaridine (Termination and extraction with NaHCO(3)
CC and ethylacetate, respectively) {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC KM=2.1 uM for salutaridine (Termination with methanol without
CC extraction) {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC KM=1.5 uM for salutaridinol {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC KM=3.5 uM for NADPH {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC KM=1190 uM for NADH {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC KM=7 uM for NADP(+) {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC Vmax=63.2 nmol/sec/mg enzyme with salutaridine as substrate
CC (Termination and extraction with NaHCO(3) and ethylacetate,
CC respectively) {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC Vmax=35.6 nmol/sec/mg enzyme with salutaridine as substrate
CC (Termination with methanol without extraction)
CC {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC Vmax=588 nmol/sec/mg enzyme with salutaridinol as substrate
CC {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC Vmax=57.3 nmol/sec/mg enzyme with NADPH as substrate
CC {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC Vmax=102 nmol/sec/mg enzyme with NADH as substrate
CC {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC Vmax=598 nmol/sec/mg enzyme with NADP(+) as substrate
CC {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC Note=The enzyme does not obey Michaelis-Menten kinetics because the
CC theorectical Vmax cannot be achieved due to strong substrate
CC inhibition, instead the optimal velocity, Vopt, is taken as the
CC measure of enzyme performance. {ECO:0000269|PubMed:17337529,
CC ECO:0000269|PubMed:19648114};
CC pH dependence:
CC Optimum pH is 6 for the reverse reaction (reduction) with activity
CC decreasing sharply towards pH 4.5 and pH 7.5. Optimum pH is 9.5 for
CC the forward reaction (oxidation) with greatly reduced activity at pH
CC 6. {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC Temperature dependence:
CC Optimum temperature is 40 degrees Celsius.
CC {ECO:0000269|PubMed:17337529, ECO:0000269|PubMed:19648114};
CC -!- BIOTECHNOLOGY: Has potential use in industrialized synthesis of the
CC antitussivum codeine and the analgesic morphine.
CC {ECO:0000269|PubMed:19648114}.
CC -!- SIMILARITY: Belongs to the short-chain dehydrogenases/reductases (SDR)
CC family. {ECO:0000255}.
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DR EMBL; EF184229; ABO93462.1; -; mRNA.
DR AlphaFoldDB; A4UHT7; -.
DR SMR; A4UHT7; -.
DR BRENDA; 1.1.1.248; 4514.
DR PRO; PR:A4UHT7; -.
DR GO; GO:0047037; F:salutaridine reductase (NADPH) activity; IEA:UniProtKB-EC.
DR GO; GO:0009820; P:alkaloid metabolic process; IEA:UniProtKB-KW.
DR CDD; cd05324; carb_red_PTCR-like_SDR_c; 1.
DR InterPro; IPR045313; CBR1-like.
DR InterPro; IPR036291; NAD(P)-bd_dom_sf.
DR InterPro; IPR002347; SDR_fam.
DR Pfam; PF00106; adh_short; 1.
DR PRINTS; PR00081; GDHRDH.
DR PRINTS; PR00080; SDRFAMILY.
DR SUPFAM; SSF51735; SSF51735; 1.
PE 1: Evidence at protein level;
KW Alkaloid metabolism; NAD; NADP; Oxidoreductase.
FT CHAIN 1..311
FT /note="Salutaridine reductase"
FT /id="PRO_0000391437"
FT ACT_SITE 236
FT /note="Proton acceptor"
FT /evidence="ECO:0000250"
FT BINDING 17..40
FT /ligand="NADP(+)"
FT /ligand_id="ChEBI:CHEBI:58349"
FT /evidence="ECO:0000250"
FT BINDING 180
FT /ligand="substrate"
FT /evidence="ECO:0000250"
FT MUTAGEN 44
FT /note="R->E: More than 50-fold decrease in affinity towards
FT NADPH."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 48
FT /note="R->E: 30-fold decrease in affinity towards NADPH.
FT 1.5-fold increase in affinity towards NADH."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 48
FT /note="R->K: 2.5-fold decrease in affinity towards NADPH.
FT 2-fold increase in affinity towards NADH."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 104
FT /note="F->A: Strong decrease in substrate affinity and
FT catalytic efficiency. Strongly reduced substrate
FT inhibition. Almost 200-fold decrease in substrate affinity
FT and 112-fold decrease in catalytic efficiency but no
FT substrate inhibition; when associated with A-275."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 106
FT /note="V->A: Decrease in substrate affinity. Increase in
FT catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 107
FT /note="D->A: 7-fold decrease in substrate affinity. 15-fold
FT decrease in catalytic efficiency. Strongly reduced
FT substrate inhibition."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 152
FT /note="N->A: 6-fold decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 180
FT /note="S->A: 3-fold decrease in substrate affinity. More
FT than 2300-fold decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 181
FT /note="S->A: More than 2-fold decrease in substrate
FT affinity. 1.5-fold decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 182
FT /note="T->A: 2-fold decrease in substrate affinity. 20-fold
FT decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 185
FT /note="L->A: 6-fold decrease in substrate affinity. 550-
FT fold decrease in catalytic efficiency. No substrate
FT inhibition."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 185
FT /note="L->S: Almost 2-fold increase in substrate affinity.
FT Almost 5-fold decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 185
FT /note="L->V: Almost 2-fold decrease in catalytic
FT efficiency."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 186
FT /note="K->V: Almost 4-fold decrease in substrate affinity.
FT More than 4-fold decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 236
FT /note="Y->F: Inactive."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 240
FT /note="K->E: Inactive."
FT /evidence="ECO:0000269|PubMed:17337529"
FT MUTAGEN 266
FT /note="L->A: Strong decrease in substrate affinity and
FT catalytic efficiency. No substrate inhibition."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 266
FT /note="L->S: 12-fold decrease in substrate affinity and 26-
FT fold decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 266
FT /note="L->V: 6-fold decrease in substrate affinity. 7-fold
FT decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 271
FT /note="M->A: Almost 11-fold decrease in substrate affinity
FT and 2200-fold decrease in catalytic efficiency but no
FT substrate inhibition."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 271
FT /note="M->T: Inactive."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 272
FT /note="N->A: Almost 43-fold decrease in substrate affinity
FT and 3300-fold decrease in catalytic efficiency but no
FT substrate inhibition."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 272
FT /note="N->T: Inactive."
FT /evidence="ECO:0000269|PubMed:17337529,
FT ECO:0000269|PubMed:19648114"
FT MUTAGEN 275
FT /note="I->A: 15-fold decrease in substrate affinity. Almost
FT 7-fold decrease in catalytic efficiency and strongly
FT reduced substrate inhibition. Almost 200-fold decrease in
FT substrate affinity and 112-fold decrease in catalytic
FT efficiency but no substrate inhibition; when associated
FT with A-104."
FT /evidence="ECO:0000269|PubMed:19648114"
FT MUTAGEN 275
FT /note="I->V: Slight decrease in catalytic efficiency."
FT /evidence="ECO:0000269|PubMed:19648114"
SQ SEQUENCE 311 AA; 34055 MW; FC0982A8160AF636 CRC64;
MPETCPNTVT KMRCAVVTGG NKGIGFEICK QLSSSGIMVV LTCRDVTRGL EAVEKLKNSN
HENVVFHQLD VTDPITTMSS LADFIKARFG KLDILVNNAG VAGFSVDADR FKAMISDIGE
DSEEVVKIYE KPEAQELMSE TYELAEECLK INYYGVKSVT EVLLPLLQLS DSPRIVNVSS
STGSLKYVSN ETALEILGDG DALTEERIDM VVNMLLKDFK ENLIETNGWP SFGAAYTTSK
ACLNAYTRVL AKKIPKFQVN CVCPGLVKTE MNYGIGNYTA DEGAKHVVRI ALFPDDGPSG
FFYDCSELSA F
