Amino acid racemase from Pseudomonas putida DSM 3263

Enzyme Description

Species

Extremophile

EC Number

5.1.1.10 ↗

Sequence

Length: 409 amino acids

MQFSRTLLAASLALLITGQAPLYAAPPLSMDNGTTALTAQNSNAWVEISAGALQHNIRTLQAELGGKSKLCAVLKADAYGHGIGLVMPSIIAQGVPCVAVASNEEARVVRASGFTGQLVRVRLASLGEVEDALQYDMEELVGSAEFARQLDAIAERHGKTLRIHMALNSSGMSRNGVEMTTWSGRGEALQITDQKHLQLVALMTHFAVEDKDDVRKGLAAFNEQTDWLIKHAKLDRSKLTLHAANSFATLEVPEAHLDMVRTGGALFGDTVPTRTEYQRVMQFKSHVAAVHSYPAGNTVGYDRTFTLARDSRLANITVGTSDGYRRVFTNKGHVLINGHRVPVVGKVSMNTLMVDVTDFPDVKGGNEVVLFGKQAGREITQAEIEEINGALLADLYTVWGSSNPKILVD

Christian Femmer et al. (2020) — Semi-rational engineering of an amino acid racemase that is stabilized in aqueous/organic solvent mixtures

Biotechnology and Bioengineering · doi:10.1002/bit.27449 ↗ · Activity - Michaelis-Menten Stability - Half-life

54 measurements ▼

Database ID

—

Sequence Annotation

Explicit - Provided

Protein Source

Recombinant, host bacterium Escherichia coli GFP JW2786-1

Experimental Data (54 measurements)

54 measurements — page 1 of 3

Mutation	Property	Assay	Solvent	Solvent Volume	Aqueous Reference	WT Reference	Measured Value	Units	Solution	pH	Temperature	Substrate(s)	Product(s)	Cofactor(s)	Shaking	Comments
WT	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	325.0	—	170.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
WT	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	40%	325.0	—	86.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
WT	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	1.3	—	3.3	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
WT	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	1.3	—	1.1	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
WT	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	40%	1.3	—	1.2	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
WT	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	325.0	—	217.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	369.0	217.0	249.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	0.9	3.3	0.7	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	369.0	170.0	218.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	40%	369.0	86.0	96.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	0.9	1.1	3.7	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	40%	0.9	1.2	1.8	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	352.0	217.0	248.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	352.0	170.0	210.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	40%	352.0	86.0	91.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	1.0	3.3	0.7	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	1.0	1.1	1.4	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)	Activity - Michaelis-Menten	Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	40%	1.0	1.2	0.6	mM	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in mM)
A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	10%	367.0	217.0	232.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)
A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)	Activity - Michaelis-Menten	kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent	Methanol	20%	367.0	170.0	199.0	1/s	20 mM Tris-HCl buffer	7	25°C	D-ornithine	L-ornithine	900 rpm	900 rpm	Classical aqueous control (in 1/s)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

325.0

—

170.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

325.0

—

86.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

1.3

—

3.3

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

1.3

—

1.1

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

1.3

—

1.2

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

325.0

—

217.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

369.0

217.0

249.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

0.9

3.3

0.7

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

369.0

170.0

218.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

369.0

86.0

96.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

0.9

1.1

3.7

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

0.9

1.2

1.8

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

352.0

217.0

248.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

352.0

170.0

210.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

352.0

86.0

91.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

1.0

3.3

0.7

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

1.0

1.1

1.4

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

1.0

1.2

0.6

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

367.0

217.0

232.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

367.0

170.0

199.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

367.0

86.0

92.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

1.0

3.3

0.7

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

1.0

1.1

4.2

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

1.0

1.2

1.4

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

0.9

1.1

2.4

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

345.0

217.0

249.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

345.0

170.0

216.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

345.0

86.0

93.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

0.9

3.3

0.6

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

0.9

1.1

1.8

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

0.9

1.2

0.6

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

350.0

217.0

242.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

350.0

170.0

194.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

0.9

3.3

0.6

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

0.9

1.1

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

0.9

1.2

0.6

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

362.0

217.0

247.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

20%

362.0

170.0

205.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

362.0

86.0

90.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

10%

0.9

3.3

0.4

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Activity - Michaelis-Menten

Km (in mM) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

0.9

1.2

2.0

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in mM)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Activity - Michaelis-Menten

kcat (in 1/s) measured by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

350.0

86.0

88.0

1/s

20 mM Tris-HCl buffer

25°C

D-ornithine

L-ornithine

900 rpm

Classical aqueous control (in 1/s)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Acetonitrile

30%

—

35.0

159.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Acetonitrile

30%

—

35.0

243.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Acetonitrile

30%

—

35.0

298.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Acetonitrile

30%

—

35.0

231.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Acetonitrile

30%

—

35.0

220.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Acetonitrile

30%

—

35.0

206.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

—

19.0

87.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

—

19.0

137.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

—

19.0

157.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

—

19.0

140.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

—

19.0

142.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4)

Stability - Half-life

Half-life (in minutes), measured by activity assay by HPLC (D-ornithine production measurement) in the presence of organic solvent

Methanol

40%

—

19.0

144.0

minutes

Incubation: 20 mM Tris-HCl buffer, Assay: 20 mM Tris-HCl

Incubation: 7, Assay: 7

Incubation: 25°C, Assay: 25°C

40 mM D-ornithine

L-ornithine

Incubation: 800 rpm, Assay: 800 rpm

Classical aqueous control (in minutes)

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Mutations in this dataset (7)

WT A62R/K159R/W182D/K211R/A220S/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9) A62R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (P3B4) K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R (P3B8) K159R/W182D/K230R/K233R/T262V/Q278R/A308R/N337R/K373R (R9F1) W182D/K230R/K233R/T262V/Q278R/A308R/N337R (R2H4) W182D/T262V/Q278R/A308R/N337R/K373R (P1G4)

Visualization : Activity — Michaelis-Menten

One bar per measurement. Colour = solvent, shade = solvent volume. — — — Reference value. Hover for details.

Mutation Effect — Michaelis-Menten

Mutation impact on kinetic parameters in the presence of organic solvent: comparison of wild-type and mutant values in identical conditions.

Visualization : Stability — Half-life

One bar per measurement. Colour = solvent, shade = solvent volume. — — — Reference value. Hover for details.

Mutation Effect

Mutation impact on enzyme stability and function in the presence of organic solvent: comparison of wild-type and mutant values in identical conditions.

Structure

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Amino acid racemase from Pseudomonas putida DSM 3263

Enzyme Description

Sequence

Experimental Data (54 measurements)

Visualization : Activity — Michaelis-Menten

Mutation Effect — Michaelis-Menten

Visualization : Stability — Half-life

Mutation Effect

Structure

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