3.4.21.62: Subtilisin
This is an abbreviated version!
For detailed information about Subtilisin, go to the full flat file.
Word Map on EC 3.4.21.62
-
3.4.21.62
-
hydrolysates
-
chymotrypsin
-
prion
-
elastase
-
papain
-
sds
-
aeruginosa
-
prpsc
-
proteinases
-
casein
-
staphylococcus
-
proprotein
-
pronase
-
convertase
-
alpha-chymotrypsin
-
scrapie
-
dnase
-
ultrafiltration
-
proteinaceous
-
spongiform
-
thermolysin
-
pmsf
-
bacteriocins
-
licheniformis
-
propeptide
-
paraffin-embedded
-
gelatin
-
protease-resistant
-
formalin-fixed
-
pcsk9
-
alpha-amylase
-
cytotoxin
-
boil
-
creutzfeldt-jakob
-
phenylmethylsulfonyl
-
amyloliquefaciens
-
pumilus
-
bromelain
-
alkaliphilic
-
shiga
-
i-converting
-
transesterification
-
furin
-
album
-
defatted
-
prodomain
-
pancreatin
-
p-nitroanilide
-
kazal
-
detergent
-
industry
-
biotechnology
-
food industry
-
medicine
-
synthesis
-
degradation
-
analysis
- 3.4.21.62
- hydrolysates
- chymotrypsin
- prion
- elastase
- papain
- sds
- aeruginosa
- prpsc
- proteinases
- casein
- staphylococcus
- proprotein
- pronase
-
convertase
- alpha-chymotrypsin
- scrapie
- dnase
-
ultrafiltration
-
proteinaceous
-
spongiform
- thermolysin
- pmsf
-
bacteriocins
- licheniformis
- propeptide
-
paraffin-embedded
- gelatin
-
protease-resistant
-
formalin-fixed
- pcsk9
- alpha-amylase
-
cytotoxin
-
boil
- creutzfeldt-jakob
-
phenylmethylsulfonyl
- amyloliquefaciens
- pumilus
- bromelain
-
alkaliphilic
-
shiga
-
i-converting
-
transesterification
- furin
- album
-
defatted
- prodomain
- pancreatin
- p-nitroanilide
-
kazal
- detergent
- industry
- biotechnology
- food industry
- medicine
- synthesis
- degradation
- analysis
Reaction
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides =
Synonyms
AcpII, Ak.1 protease, Alcalase, Alcalase 0.6L, Alcalase 2.5L, ALE1 subtilase, ALK-enzyme, Alkaline mesentericopeptidase, Alkaline protease, alkaline serine protease, ALP1, Alzwiprase, aprE, AprE51, aqualysin, Arp, AsES, Asp v 13, AtSBT1.9, Bacillopeptidase A, Bacillopeptidase B, Bacillus gibsonii alkaline protease, Bacillus subtilis alkaline proteinase Bioprase, BgAP, Bioprase AL 15, Bioprase APL 30, BLS, BPN', BprB, BprV, C1 subtilase, cold active subtilisin-like serine proteinase, Colistinase, EC 3.4.21.14, EC 3.4.4.16, Esperase, Fe protease, Fe prtS8A, Genenase I, intracellular subtilisin protease, ISP, IvaP, Kazusase, Maxatase, mesenteroicopeptidase, More, Nagarse, Opticlean, ORF2, Orientase 10B, P69 subtilase, PBANKA_1106900, PbSOPT, Peptidase, subtilo-, A, PF3D7_0507300, phytophase, PIMMS2, Protease S, Protease VIII, Protease XXVII, Proteinase K, Proteinase, Bacillus subtilis alkaline, Protin A 3L, PSP-3, psychrophilic subtilisin-like protease, S1P subtilase, SAP, SAS-1, SASP, saspase, Savinase, Savinase 16.0L, Savinase 32.0 L EX, Savinase 4.0T, Savinase 8.0L, savinaseTM, SBc, SBL, SDD1 subtilase, senescence-associated subtilisin protease, SES7, SISBT3 subtilase, SOPT, SP 266, Sspa, SSU0757, SUB1, SUB2, subC, subtilase, subtilase subfamily 1 member 9, subtilase-like protease, subtilisin, subtilisin 72, subtilisin A, Subtilisin amylosacchariticus, Subtilisin BL, subtilisin BPN’, subtilisin C., subtilisin Carlsberg, subtilisin DJ-4, Subtilisin DY, Subtilisin E, subtilisin E-S7, Subtilisin GX, subtilisin JB1, subtilisin Karlsberg, Subtilisin Novo, subtilisin Pr1-like protease, subtilisin protease, subtilisin QK, Subtilisin S41, subtilisin S4I, subtilisin S88, Subtilisin Sendai, subtilisin Sph, subtilisin-like ookinete protein, subtilisin-like ookinete protein important for transmission, subtilisin-like protease, subtilisin-like protease AprV2, subtilisin-like serine protease, Subtilisn J, Subtilopeptidase, Superase, thermitase, thermo-active subtilisin-like serine protease, Thermoase, Thermoase PC 10, thermophilic thermitase, thermostable subtilisin, ThSS45, Tk-subtilisin, trans-cinnamoyl-subtilisin, V. cholerae-secreted serine protease, VC_0157, vPR
ECTree
3.4.21.62 SubtilisinAdvanced search results
results (
results (Engineering
Engineering on EC 3.4.21.62 - Subtilisin
Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
top
PROTEIN VARIANTS 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
N154D
GN111900.1
site-directed mutagenesis, the mutant has 107% proteolytc activity compared to the wild-type enzyme
N167D
GN111900.1
site-directed mutagenesis, the mutant has 77% proteolytc activity compared to the wild-type enzyme
N205D
GN111900.1
site-directed mutagenesis, the mutant has 113% proteolytc activity compared to the wild-type enzyme
N236D
GN111900.1
site-directed mutagenesis, the mutant has 115% proteolytc activity compared to the wild-type enzyme
N242D
GN111900.1
site-directed mutagenesis, the mutant has 118% proteolytc activity compared to the wild-type enzyme
N250D
GN111900.1
site-directed mutagenesis, the mutant has 113% proteolytc activity compared to the wild-type enzyme
N253D
GN111900.1
site-directed mutagenesis, the mutant has 135% proteolytc activity compared to the wild-type enzyme and shows a dramatic pH activity profiles shifted towards higher activity at lower pH range of pH 8.5-10
N253D/Q256E
GN111900.1
site-directed mutagenesis, the mutant shows a dramatic pH activity profiles shifted towards higher activity at lower pH range of pH 8.5-10, the mutant has a 2fold increased activity compared to the wild-type enzyme with a thermal resistance increased by 2.4°C at pH 8.5
N60D
GN111900.1
site-directed mutagenesis, the mutant has 32% proteolytc activity compared to the wild-type enzyme
N97D
GN111900.1
site-directed mutagenesis, the mutant has 146% proteolytc activity compared to the wild-type enzyme
Q107E
GN111900.1
site-directed mutagenesis, the mutant has 53% proteolytc activity compared to the wild-type enzyme
Q12E
GN111900.1
site-directed mutagenesis, the mutant has 114% proteolytc activity compared to the wild-type enzyme
Q176E
GN111900.1
site-directed mutagenesis, the mutant has 97% proteolytc activity compared to the wild-type enzyme
Q200E
GN111900.1
site-directed mutagenesis, the mutant has 125% proteolytc activity compared to the wild-type enzyme
Q230E
GN111900.1
site-directed mutagenesis, the mutant has 107% proteolytc activity compared to the wild-type enzyme
Q256E
GN111900.1
site-directed mutagenesis, the mutant has 129% proteolytc activity compared to the wild-type enzyme and shows a dramatic pH activity profiles shifted towards higher activity at lower pH range of pH 8.5-10
Q37E
GN111900.1
site-directed mutagenesis, the mutant has 136% proteolytc activity compared to the wild-type enzyme
N250D
Alkalihalobacillus gibsonii DSM 14391
-
site-directed mutagenesis, the mutant has 113% proteolytc activity compared to the wild-type enzyme
-
N97D
Alkalihalobacillus gibsonii DSM 14391
-
site-directed mutagenesis, the mutant has 146% proteolytc activity compared to the wild-type enzyme
-
Q107E
Alkalihalobacillus gibsonii DSM 14391
-
site-directed mutagenesis, the mutant has 53% proteolytc activity compared to the wild-type enzyme
-
Q12E
Alkalihalobacillus gibsonii DSM 14391
-
site-directed mutagenesis, the mutant has 114% proteolytc activity compared to the wild-type enzyme
-
A1C
does not change the catalytic properties of subtilisin S189 but allows the introduction of a fluorescent group at its N-terminus
D32A
engineered variant of the protease subtilisin, denoted S189, mutation renders the enzyme's activity dependent on the presence of certain small anions such as fluoride or azide. Is activated more than 3000fold by azide
G169A
L126I
M50F
N218S
N76D
Q271E
S101W/G169A/V192A
-
mutant shows 1.4fold activity compared to wild-type, kcat doubled compared to wild-type, Km decreased compared to wild-type
S188P
S194P
S221A
S221C
T22C/S87C/S221C
-
if denatured, it is refolded with an excess of proR9. About 75% is rapidly bound to proR9. 25% undergoes a slow step prior to binding
T254A
Y217K
G165L
site-directed mutagenesis, the mutant shows slightly increased transacylation activity compared to wild-type
G165L/M221C
site-directed mutagenesis, the mutant shows about 5fold increased transacylation activity compared to wild-type
G165L/M221F
site-directed mutagenesis, the mutant shows about 6.5fold increased transacylation activity compared to wild-type
G165L/M221S
site-directed mutagenesis, the mutant shows slightly increased transacylation activity compared to wild-type
G165X
site-directed mutagenesis, catalytic constants of subtilisin Carlsberg for perhydrolysis of methylpropionate, methylbutyrate and methylpentanoate are increased up to 3.5fold, 5.4fold and 5.5fold, respectively, while proteolysis is decreased up to 100fold for N-succinyl-Ala-Ala-Pro-Phe-pnitroanilide substrate
G166R
-
the mutant is both more stable and displays more activity compared to the wild-type enzyme
M221A
site-directed mutagenesis, the mutant shows reduced transacylation activity compared to wild-type
M221C
site-directed mutagenesis, the mutant shows similar transacylation activity compared to wild-type
M221F
site-directed mutagenesis, the mutant shows similar transacylation activity compared to wild-type
M221S
site-directed mutagenesis, the mutant shows unaltered transacylation activity compared to wild-type
M221W
site-directed mutagenesis, the mutant shows reduced transacylation activity compared to wild-type
N212G
site-directed mutagenesis, the mutant shows increased kcat compared to the wild-type enzyme
P210A
site-directed mutagenesis, the mutant shows increased kcat compared to the wild-type enzyme
P210G
site-directed mutagenesis, the mutant shows 1.5fold increased kcat compared to the wild-type enzyme
P210G/T211G
site-directed mutagenesis, the mutant shows 1.7fold increased kcat compared to the wild-type enzyme
P210G/T211G/N212G
site-directed mutagenesis, the mutant shows 1.8fold increased kcat compared to the wild-type enzyme
S161C
-
the mutant brings increased stability to subtilisin compared to the wild-type
T211G
site-directed mutagenesis, the mutant shows 1.8fold increased kcat compared to the wild-type enzyme
T58A/L216W
site-directed mutagenesis, the mutant catalyzes in addition to its proteolytic activity the generation of peroxycarboxylic acids from corresponding esters in the presence of hydrogen peroxide
A153V
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, but does not show improved properties in chaotropic conditions
G166M
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, and improved properties in chaotropic conditions
G166S
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, and improved properties in chaotropic conditions
I205V
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, but does not show improved properties in chaotropic conditions
K211P/R212A
mutant is more thermostable compared to wild-type, half-life at 60°C 10times longer compared to wild-type. Molecular dynamics simulation at 10°C and 90°C reveal that the average global flexibility of both variants is slightly higher than wild-type
K211P/R212A/S145I/S175T/K221E/N291I/S295T
mutant is more thermostable compared to wild-type, half-life at 60°C 500times longer compared to wild-type. Molecular dynamics simulation at 10°C and 90°C reveal that the average global flexibility of both variants is slightly higher than wild-type
N218S
site-directed mutagenesis, the mutant shows increased activity and stability in the presence of GdmCl or SDS compared to the wild-type enzyme
Q125/Q377E/Q381R
site-directed mutagenesis, the mutant shows increased thermotolerance compared to the wild-type enzyme, but is 60% less active than the wild-type
Q125R
site-directed mutagenesis, the mutant does not show increased thermotolerance compared to the wild-type enzyme, it is 60% less active than the wild-type
Q377E
site-directed mutagenesis, the mutant shows increased thermotolerance and an increase in protease activity of 46.5% compared to the wild-type enzyme
Q381R
site-directed mutagenesis, the mutant shows increased thermotolerance and an increase in protease activity of 46.5% compared to the wild-type enzyme
S221C
-
site-directed mutagenesis, mutation of the critical catalytic residue Ser221 of SES7 to a cysteine to avoid protease self-digestion
S62I
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, and slightly improved properties in chaotropic conditions
S62I/A153V/G166S/I205V
S62I/A153V/G166S/I205V/N218S/T224A
site-directed mutagenesis, the mutant shows high activity and stability in the presence of GdmCl or SDS compared to the wild-type enzyme
S62I/A153V/G166S/T224A/T240S
S62I/G166M
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, and highly improved properties in chaotropic conditions
T224A
site-directed mutagenesis, the mutant shows increased activity and stability in the presence of GdmCl or SDS compared to the wild-type enzyme
Q125/Q377E/Q381R
-
site-directed mutagenesis, the mutant shows increased thermotolerance compared to the wild-type enzyme, but is 60% less active than the wild-type
-
Q125R
-
site-directed mutagenesis, the mutant does not show increased thermotolerance compared to the wild-type enzyme, it is 60% less active than the wild-type
-
Q377E
-
site-directed mutagenesis, the mutant shows increased thermotolerance and an increase in protease activity of 46.5% compared to the wild-type enzyme
-
Q381R
-
site-directed mutagenesis, the mutant shows increased thermotolerance and an increase in protease activity of 46.5% compared to the wild-type enzyme
-
S221C
Bacillus subtilis JN-S7
-
site-directed mutagenesis, mutation of the critical catalytic residue Ser221 of SES7 to a cysteine to avoid protease self-digestion
-
D180G
-
mutant using substrate N-methoxysuccinyl-Ala-Ala-Pro-Val-4-nitroanilide Km similar to wild-type, kcat value enhanced, kcat/Km value enhanced by 4%
D180G/D182G
-
using substrate N-methoxysuccinyl-Ala-Ala-Pro-Val-4-nitroanilide Km similar to wild-type, kcat value highly enhanced, kcat/Km value enhanced by 33%
D182G
-
using substrate N-methoxysuccinyl-Ala-Ala-Pro-Val-4-nitroanilide Km similar to wild-type, kcat value enhanced, kcat/Km value enhanced by 27%
S166C
-
site-directed mutagenesis combined with chemical modification
V104S/L124M/P129S/S130G/P131E/A133S/T134L/L135I
-
mutant containing the highly flexible region of psychrophilic enzyme TA39 subtilisin. Mutant shows the same temperature optimum and pH-profile as wild-type, but higher specificity for substrate succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide and broader substrate specificity. Mutant has a decreased thermostability, but increased activity at low temperature
G6G
-
possesses a single Tn917 insertion, is devoid of subtilisin-like proteinase activity, has longer generation times and is more susceptible to killing by whole blood than the wild-type
M3G
-
possesses a single Tn917 insertion, is devoid of subtilisin-like proteinase activity, has longer generation times and is more susceptible to killing by whole blood than the wild-type
G6G
-
possesses a single Tn917 insertion, is devoid of subtilisin-like proteinase activity, has longer generation times and is more susceptible to killing by whole blood than the wild-type
-
M3G
-
possesses a single Tn917 insertion, is devoid of subtilisin-like proteinase activity, has longer generation times and is more susceptible to killing by whole blood than the wild-type
-
DELTACa2-Pro-S324A
-
does not completely lose the ability to fold into a native structure, probably because binding of the water molecule to the position corresponding to the Ca2 site and movement of the epsilon-amino group of Lys213 to the position corresponding to the Ca3 site compensate at least partly for the destabilization of the structure of the Ca2+-binding loop caused by the removal of the Ca2 or Ca3 site
DELTACa3-Pro-S324A
-
does not completely lose the ability to fold into a native structure, probably because binding of the water molecule to the position corresponding to the Ca2 site and movement of the epsilon-amino group of Lys213 to the position corresponding to the Ca3 site compensate at least partly for the destabilization of the structure of the Ca2+-binding loop caused by the removal of the Ca2 or Ca3 site
DELTAloop-subtilisin
-
the unique insertion sequence of subtilisin (Pro207-Asp226) is removed and Gly206,Ala228, and Glu229 are replaced with Asn, Met, and Asp, respectively. Lacks the Ca2+-binding loop, completely loses the ability to fold into a native structure
E61G
-
pro-subtilisin mutant, exhibits a halo-forming activity at 80, 70 and 60°C
E61K
-
pro-subtilisin mutant, exhibits a halo-forming activity at 80, 70 and 60°C
G56E
-
Pro-G56E gives a halo at temperatures equal or higher than 60 °C
G56F
-
Pro-G56F gives a halo only at temperatures equal or higher than 70 °C
G56S
G56S/T135S
-
low-temperature adaptation of prepro-G56S/T135S-subtilisin
S255A
-
active-site mutant of pro-subtilisin. Accumulates in cells in inclusion bodies like pro-subtilisin
S324A
S359A
-
active site mutant. Upon overproduction, Pro-S359A accumulates in the cells in a soluble form
S361A
additional information
L126I
-
site-directed mutagenesis, BPN', stable only in presence of excess Ca2+
M50F
-
site-directed mutagenesis, BPN',stabilized in EDTA, but destabilized slightly in CaCl2
N218S
-
site-directed mutagenesis, BPN', stabilized significantly, either with or without Ca2+
Q271E
-
site-directed mutagenesis, BPN', stabilized significantly, either with or without Ca2+
S188P
-
site-directed mutagenesis, BPN', stable only in presence of excess Ca2+
S221A
-
a delta75-83 mutant. Shows slow folding reaction. Is unstable at low metal concentration, unfolding at a rate of 0.6 h-1. If denatured, it is refolded with an excess of proR9
T254A
-
site-directed mutagenesis, BPN', stable only in presence of excess Ca2+
Y217K
-
site-directed mutagenesis, BPN', stabilized significantly, either with or without Ca2+
S62I/A153V/G166S/I205V
site-directed mutagenesis, the mutant shows increased activity and stability in the presence of GdmCl or SDS compared to the wild-type enzyme
S62I/A153V/G166S/I205V
site-directed mutagenesis, the mutant shows increased activity without detergents compared to the wild-type, and highly improved properties in chaotropic conditions
S62I/A153V/G166S/T224A/T240S
site-directed mutagenesis, the mutant shows high activity and stability in the presence of GdmCl or SDS compared to the wild-type enzyme
S62I/A153V/G166S/T224A/T240S
site-directed mutagenesis, the mutant shows increased activity and stability in the presence of GdmCl or SDS compared to the wild-type enzyme
G56S
-
pro-subtilisin mutant, does not seriously affect the structure of pro-subtilisin. Is more effectively matured than pro-subtilisin at lower temperatures
S324A
-
pro-subtilisin, which is not autoprocessed at all when the active site serine residue is replaced with Ala. Is refolded in the presence of Ca2+
S324A
-
Pro-S324A represents the active site mutant of pro-subtilisin, Pro-S324A is used to construct mutant proteins to prevent their autoprocessing and self-degradation
S361A
site-directed mutagenesis, mutation of the catalytic residue partially inhibits autoproteolysis, but the IvaPS361A mutant can be cleaved to the 47-kDa mature form through a mechanism other than autoproteolysis
S361A
-
site-directed mutagenesis, mutation of the catalytic residue partially inhibits autoproteolysis, but the IvaPS361A mutant can be cleaved to the 47-kDa mature form through a mechanism other than autoproteolysis
-
S361A
Vibrio cholerae serotype O1 C6706
-
site-directed mutagenesis, mutation of the catalytic residue partially inhibits autoproteolysis, but the IvaPS361A mutant can be cleaved to the 47-kDa mature form through a mechanism other than autoproteolysis
-
S361A
-
site-directed mutagenesis, mutation of the catalytic residue partially inhibits autoproteolysis, but the IvaPS361A mutant can be cleaved to the 47-kDa mature form through a mechanism other than autoproteolysis
-
additional information
GN111900.1
mimicking surface deamidation by substituting Gln by Glu and/or Asn by Asp might be a simple and fast protein reengineering approach for modulating enzyme properties such as activity, pH optimum, and thermal resistance, deamidation in five (N97, N253, Q37, Q200, and Q256) out of eight (N97, N154, N250, N253, Q37, Q107, Q200, and Q256) amino acids, comparison of wild-type and mutant enzymes, overview
additional information
-
mimicking surface deamidation by substituting Gln by Glu and/or Asn by Asp might be a simple and fast protein reengineering approach for modulating enzyme properties such as activity, pH optimum, and thermal resistance, deamidation in five (N97, N253, Q37, Q200, and Q256) out of eight (N97, N154, N250, N253, Q37, Q107, Q200, and Q256) amino acids, comparison of wild-type and mutant enzymes, overview
additional information
Alkalihalobacillus gibsonii DSM 14391
-
mimicking surface deamidation by substituting Gln by Glu and/or Asn by Asp might be a simple and fast protein reengineering approach for modulating enzyme properties such as activity, pH optimum, and thermal resistance, deamidation in five (N97, N253, Q37, Q200, and Q256) out of eight (N97, N154, N250, N253, Q37, Q107, Q200, and Q256) amino acids, comparison of wild-type and mutant enzymes, overview
-
additional information
-
introduction of about 20 site-directed mutations to obtain a variant with increased sequence selectivity, a predomain that can direct cleavage to the junction of any protein fused to it, and a modified active site to kinetically isolate binding and cleavage reactions. Identification of specific anions that trigger the processing reaction so that column-immobilized mutant enzyme can be used as both the affinity ligand and the processing protease for one-step purification methods
additional information
-
sequential randomization of 12 amino acid positions in calcium-free enzyme. Mutations increase the half-life of enzyme at elevated temperature by 15000fold and partially increase ratio of kcat/KM-value
additional information
site saturation mutagenesis of the subtilisin Carlsberg variant T58A/L216W at positionsThr33, Val67, Ser124, Gly126, Ser155, Gly165, Ala168, Trp216, Asn217, Gly218, Met221
additional information
immobilization of recombinant subtilisin Carlsberg. Enantioselectivities of immobilized wild-type and mutants of subtilisin Carlsberg in the transacylation of racemic 1-phenylethanol and vinyl butyrate in tetrahydrofuran
additional information
-
ionic-surfactant-coated Bacillus licheniformis subtilisin (ISCBLS) is prepared by freeze-drying Bacillus licheniformis subtilisin with both ionic surfactant and dextrin. ISCBCL displays 9300fold enhanced activity relative to its native counterpart in the transesterificaion of N-acetyl phenylalanine ethyl ester with 1-propanol in hexane and 12800fold enhanced activity in the transesterification of trifluoroethyl butyrate with 1-phenylethanol in tetrahydrofuran
additional information
-
the enzyme subtilisin A is immobilized by simple hydrophobic adsorption onto various surface-grafted macroporous silica gels resulting in easy-to-prepare and stable biocatalysts enabling the efficient kinetic resolution (KR) and dynamic kinetic resolution (DKR) of racemic N-tert-butyloxycarbonylphenylalanine ethyl thioester with benzylamine
additional information
-
mutant lacking three secreted serine proteases regulated by sigma-H, subtilisin, Epr and Vpr, produce less CSF and has less proCSF-processing activity
additional information
directed evolution of subtilisin E into a highly active and guanidinium chloride- and sodium dodecylsulfate-tolerant protease, subtilisin E is engineered into a chaotolerent protease, the enzyme gains a protease stability improved in a chaotropic salt (GdmCl) or a detergent (SDS), nultiple screening, overview
additional information
CRISPR-Cas9 in situ engineering of subtilisin E in Bacillus subtilis by usage of a flexible, co-transformation approach where the single guide RNA is inserted in a plasmid for Cas9 co-expression, and the donor DNA is supplied as a linear PCR product observing an editing efficiency of 76%. The method allows multiple, rapid rounds of in situ editing of the subtilisin E gene to incorporate a salt bridge triad present in the Bacillus clausii thermotolerant homologue, M-protease. Method, overview. The enzyme mutant obtained shows increased thermotolerance and activity
additional information
-
CRISPR-Cas9 in situ engineering of subtilisin E in Bacillus subtilis by usage of a flexible, co-transformation approach where the single guide RNA is inserted in a plasmid for Cas9 co-expression, and the donor DNA is supplied as a linear PCR product observing an editing efficiency of 76%. The method allows multiple, rapid rounds of in situ editing of the subtilisin E gene to incorporate a salt bridge triad present in the Bacillus clausii thermotolerant homologue, M-protease. Method, overview. The enzyme mutant obtained shows increased thermotolerance and activity
additional information
-
CRISPR-Cas9 in situ engineering of subtilisin E in Bacillus subtilis by usage of a flexible, co-transformation approach where the single guide RNA is inserted in a plasmid for Cas9 co-expression, and the donor DNA is supplied as a linear PCR product observing an editing efficiency of 76%. The method allows multiple, rapid rounds of in situ editing of the subtilisin E gene to incorporate a salt bridge triad present in the Bacillus clausii thermotolerant homologue, M-protease. Method, overview. The enzyme mutant obtained shows increased thermotolerance and activity
-
additional information
generation of enzyme-deficient mutant PbDELTASOPT/PIMMS2, production of oocysts and sporozoites in mosquitoes containing PbDELTASOPT/PIMMS2 parasites
additional information
-
generation of enzyme-deficient mutant PbDELTASOPT/PIMMS2, production of oocysts and sporozoites in mosquitoes containing PbDELTASOPT/PIMMS2 parasites
additional information
-
generation of enzyme-deficient mutant PbDELTASOPT/PIMMS2, production of oocysts and sporozoites in mosquitoes containing PbDELTASOPT/PIMMS2 parasites
-
additional information
construction of pro-enzyme derivatives lacking the Ca1 ion (Pro-TKS/DELTACa1), Ca6 ion (Pro-TKS/DELTACa6), and Ca7 ion (Pro-TKS/DELTACa7), and their active site mutants, Pro-S324A/DELTACa1, Pro-S324A/DELTACa6, and Pro-S324A/DELTACa7, respectively. Mutants Pro-TKS/DELTACa6 and Pro-TKS/DELTACa7 fully mature into their active forms upon incubation at 80°C for 30 min like the wild-type enzyme. In contrast, mutant Pro-TKS/DELTACa1 matures poorly at 80°C because of the instability of its mature domain. Refolding rates of all Pro-S324A derivatives are comparable to that of Pro-S324A (active site mutant of wild-type pro-enzyme), indicating that these Ca(2+) ions are not needed for folding of Tk-subtilisin
additional information
-
construction of pro-enzyme derivatives lacking the Ca1 ion (Pro-TKS/DELTACa1), Ca6 ion (Pro-TKS/DELTACa6), and Ca7 ion (Pro-TKS/DELTACa7), and their active site mutants, Pro-S324A/DELTACa1, Pro-S324A/DELTACa6, and Pro-S324A/DELTACa7, respectively. Mutants Pro-TKS/DELTACa6 and Pro-TKS/DELTACa7 fully mature into their active forms upon incubation at 80°C for 30 min like the wild-type enzyme. In contrast, mutant Pro-TKS/DELTACa1 matures poorly at 80°C because of the instability of its mature domain. Refolding rates of all Pro-S324A derivatives are comparable to that of Pro-S324A (active site mutant of wild-type pro-enzyme), indicating that these Ca(2+) ions are not needed for folding of Tk-subtilisin
