Did you know ? If you order before Friday 14h we deliver 90PCT of the the time next Tuesday, GENTAUR another in time delivery

Pubmed ID :23913421
Publication Date : //

Directed evolution and structural analysis of NADPH-dependent Acetoacetyl Coenzyme A (Acetoacetyl-CoA) reductase from Ralstonia eutropha reveals two mutations responsible for enhanced kinetics.


NADPH-dependent acetoacetyl-coenzyme A (acetoacetyl-CoA) reductase (PhaB) is a key enzyme in the synthesis of poly(3-hydroxybutyrate) [P(3HB)], along with β-ketothiolase (PhaA) and polyhydroxyalkanoate synthase (PhaC). In this study, PhaB from Ralstonia eutropha was engineered by means of directed evolution consisting of an error-prone PCR-mediated mutagenesis and a P(3HB) accumulation-based in vivo screening system using Escherichia coli. From approximately 20,000 mutants, we obtained two mutant candidates bearing Gln47Leu (Q47L) and Thr173Ser (T173S) substitutions. The mutants exhibited kcat values that were 2.4-fold and 3.5-fold higher than that of the wild-type enzyme, respectively. In fact, the PhaB mutants did exhibit enhanced activity and P(3HB) accumulation when expressed in recombinant Corynebacterium glutamicum. Comparative three-dimensional structural analysis of wild-type PhaB and highly active PhaB mutants revealed that the beneficial mutations affected the flexibility around the active site, which in turn played an important role in substrate recognition. Furthermore, both the kinetic analysis and crystal structure data supported the conclusion that PhaB forms a ternary complex with NADPH and acetoacetyl-CoA. These results suggest that the mutations affected the interaction with substrates, resulting in the acquirement of enhanced activity.

Authors : Matsumoto Ken'ichiro , Tanaka Yoshikazu , Watanabe Tsuyoshi , Motohashi Ren , Ikeda Koji , Tobitani Kota , Yao Min , Tanaka Isao , Taguchi Seiichi ,

Related products :

Catalog number Product name Quantity
2415-85-2 n-acetoacetyl-p-toluidine n-acetoacetyl-p-toluidin 1g
GWB-678FF0 Anti- ACAT2 (acetyl-Coenzyme A acetyltransferase 2 (acetoacetyl Coenzyme A thiolase)) Antibody
GWB-B0ACA4 Anti- ACAT2 (acetyl-Coenzyme A acetyltransferase 2 (acetoacetyl Coenzyme A thiolase)) Antibody
201-20-0127 ACAT1{Homo sapiens acetyl-Coenzyme A acetyltransferase 1 (acetoacetyl Coenzyme A thiolase),}rabbit.pAb 0.1ml
102029-52-7 Acetoacetyl coenzyme A sodium salt hyd Coenzyme A 1g
201-12-3068 Human Acetoacetyl Coenzyme A Synthetase(AACS)ELISA Kit 96T
UB-E00454 Human Acetoacetyl Coenzyme A Synthetase(AACS)ELISA Kit 96T
QY-E00454 Human Acetoacetyl Coenzyme A Synthetase(AACS)ELISA Kit 96T
26-326 This enzyme condenses acetyl-CoA with acetoacetyl-CoA to form HMG-CoA, which is the substrate for HMG-CoA reductase. 0.05 mg
29-598 HMGCS2 condenses acetyl-CoA with acetoacetyl-CoA to form HMG-CoA, which is the substrate for HMG-CoA reductase. 0.1 mg
29-599 This enzyme condenses acetyl-CoA with acetoacetyl-CoA to form HMG-CoA, which is the substrate for HMG-CoA reductase. 0.1 mg
EIAAB05623 Adipocyte protein P27,AP27,Carbonyl reductase [NADPH] 2,Cbr2,LCR,Lung carbonyl reductase,Mouse,Mus musculus,NADPH-dependent carbonyl reductase 2
3103-38-6 N-(Acetoacetyl)glycine N-(Acetoacetyl)glycine 1g
EIAAB05619 15-hydroxyprostaglandin dehydrogenase [NADP+],Bos taurus,Bovine,Carbonyl reductase [NADPH] 1,CBR1,NADPH-dependent carbonyl reductase 1,Prostaglandin 9-ketoreductase,Prostaglandin-E(2) 9-reductase
EIAAB05620 15-hydroxyprostaglandin dehydrogenase [NADP+],Carbonyl reductase [NADPH] 1,Cbr,Cbr1,Mouse,Mus musculus,NADPH-dependent carbonyl reductase 1,Prostaglandin 9-ketoreductase,Prostaglandin-E(2) 9-reductase
EIAAB05624 Carbonyl reductase [NADPH] 3,CBR3,Homo sapiens,Human,NADPH-dependent carbonyl reductase 3
EIAAB26662 Homo sapiens,Human,NADPH-dependent diflavin oxidoreductase 1,NADPH-dependent FMN and FAD-containing oxidoreductase,NDOR1,Novel reductase 1,NR1
EIAAB05622 Carbonyl reductase [NADPH] 2,CBR2,LCR,Lung carbonyl reductase,NADPH-dependent carbonyl reductase 2,Pig,Sus scrofa
EIAAB05617 15-hydroxyprostaglandin dehydrogenase [NADP+],Carbonyl reductase [NADPH] 1,CBR,CBR1,CRN,Homo sapiens,Human,NADPH-dependent carbonyl reductase 1,Prostaglandin 9-ketoreductase,Prostaglandin-E(2) 9-reduc
EIAAB05621 15-hydroxyprostaglandin dehydrogenase [NADP+],20-beta-hydroxysteroid dehydrogenase,Carbonyl reductase [NADPH] 1,CBR,CBR1,NADPH-dependent carbonyl reductase 1,Oryctolagus cuniculus,Prostaglandin 9-keto
EIAAB05616 15-hydroxyprostaglandin dehydrogenase [NADP+],20-beta-hydroxysteroid dehydrogenase,Carbonyl reductase [NADPH] 1,CBR,CBR1,CRN,NADPH-dependent carbonyl reductase 1,Pig,Prostaglandin 9-ketoreductase,Pros
EIAAB05618 15-hydroxyprostaglandin dehydrogenase [NADP+],20-beta-hydroxysteroid dehydrogenase,Carbonyl reductase [NADPH] 1,Cbr,Cbr1,NADPH-dependent carbonyl reductase 1,Prostaglandin 9-ketoreductase,Prostaglandi
EIAAB44144 NADPH-dependent thioredoxin reductase,Rat,Rattus norvegicus,Thioredoxin reductase 1, cytoplasmic,Thioredoxin reductase TR1,TR,Trxr1,Txnrd1
93-68-5 o-Acetoacetotoluidide AAOT ; N-Acetoacetyl- 1g
B11470 N n-Bis(acetoacetyl)-O-Toluidine 5 G


https://antibody-antibodies.com/ | https://gentaur.com/ | https://gen-script.com/ | https://diagenox.com/ | https://clonagen.com/ | http://gentaursearch.com/ | http://gentaurpub.com/ | https://gentaur-online.com/ | http://anti-anti-pdf.com/ | http://gentaur-worldwide.com/

 

GENTAUR Belgium BVBA BE0473327336
Voortstraat 49, 1910 Kampenhout BELGIUM
Tel 0032 16 58 90 45

Fax 0032 16 50 90 45
info@gentaur.com | Gentaur | Gentaur





GENTAUR Ltd.
Unicorn House, Station Cl
Hertfordshire, Potters Bar EN6 1TL
Whetstone London N20 9BH
Tel 020 3393 8531 Fax 020 8445 9411
uk@gentaur.com | Gentaur | Gentaur

 

 




GENTAUR France SARL
9, rue Lagrange, 75005 Paris
Tel 01 43 25 01 50

Fax 01 43 25 01 60
RCS Paris B 484 237 888

SIRET 48423788800017
RIB 30004 00187 00010092253 10
BNP PARIBAS PARIS PL MAUBERT BIC BNPAFRPPPRG
IBAN FR76 3000 4001 8700 0100 9225 310
france@gentaur.com | Gentaur | Gentaur

GENTAUR GmbH
Marienbongard 20
52062 Aachen Deutschland
Support Karolina Elandt
Tel: +49 0241 40 08 90 86, +49 0241 95 78 94 78, +49 0241 40 08 90 86
Fax: (+49) 241 56 00 47 88

Logistic :0241 40 08 90 86
Bankleitzahl 39050000
IBAN lautet DE8839050000107569353
Handelsregister Aachen HR B 16058
Umsatzsteuer-Identifikationsnummer *** DE 815175831
Steuernummer 201/5961/3925
de@gentaur.com | Gentaur | Gentaur

GENTAUR U.S.A
Genprice Inc, Logistics
547, Yurok Circle
San Jose, CA 95123
CA 95123
Tel (408) 780-0908,
Fax (408) 780-0908,
sales@genprice.com

Genprice Inc, Invoices and accounting
6017 Snell Ave, Ste 357
San Jose, CA 95123




GENTAUR Nederland BV
NL850396268B01 KVK nummer 52327027
Kuiper 1
5521 DG Eersel Nederland
Tel:  0208-080893  Fax: 0497-517897
nl@gentaur.com | Gentaur | Gentaur
IBAN: NL04 RABO 0156 9854 62   SWIFT RABONL2U






GENTAUR Spain
tel:0911876558
spain@gentaur.com | Gentaur | Gentaur






ГЕНТАУЪР БЪЛГАРИЯ
ID # 201 358 931 /BULSTAT
София 1000, ул. "Граф Игнатиев" 53 вх. В, ет. 2
Tel 0035924682280 Fax 0035924808322
e-mail: Sofia@gentaur.com | Gentaur | Gentaur
IBAN: BG11FINV91501014771636
BIC: FINVBGSF

GENTAUR Poland Sp. z o.o.


ul. Grunwaldzka 88/A m.2
81-771 Sopot, Poland
TEL Gdansk 058 710 33 44 FAX  058 710 33 48              

poland@gentaur.com | Gentaur | Gentaur

Other countries

Österreich +43720880899

Canada Montreal +15149077481

Ceská republika Praha +420246019719

Danmark +4569918806

Finland Helsset +358942419041

Magyarország Budapest +3619980547

Ireland Dublin+35316526556

Luxembourg+35220880274

Norge Oslo+4721031366

Sverige Stockholm+46852503438

Schweiz Züri+41435006251

US New York+17185132983

GENTAUR Italy
SRL IVA IT03841300167
Piazza Giacomo Matteotti, 6
24122 Bergamo Tel 02 36 00 65 93
Fax 02 36 00 65 94
italia@gentaur.com | Gentaur | Gentaur