Literature summary for 4.2.1.1 extracted from

Warden, A.C.; Williams, M.; Peat, T.S.; Seabrook, S.A.; Newman, J.; Dojchinov, G.; Haritos, V.S.
Rational engineering of a mesohalophilic carbonic anhydrase to an extreme halotolerant biocatalyst (2015), Nat. Commun., 6, 10278 .

Search for more literature for 4.2.1.1

Cloned(Commentary)

Cloned (Comment)	Organism
gene CA2, recombinant expression of wild-type and mutant enzymes in Escherichia coli	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant wild-type enzyme and mutants M1-M4, X-ray diffraction structure determination and analysis at resolutions of 1.62-2.29 A	Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
G8D/K18E/N24D/K36D/V39D/V50D/R57D/N62D/Q74E/T85D/Q136E/K169E/N177D/N186E/Q220E/L238E/N252D/Q254E	site-directed mutagenesis, mutant M4, shows reduced activity but increased thermostability and halostability compared to wild-type enzyme. The successful redesign of a mesohalophile enzyme to an extremely halotolerant orthologue, that is, one that is active at 3M NaCl and above. Role of Na+ in stabilizing M4 structure, molecular dynamics simulations, overview	Homo sapiens
G8D/K36D/V50D/N62D/Q136E/L238E	site-directed mutagenesis, mutant M2, shows reduced activity but increased thermostability and halostability compared to wild-type enzyme	Homo sapiens
G8D/N24D/K36D/V39D/V50D/R57D/N62D/Q74E/Q136E/K169E/L238E/N252D	site-directed mutagenesis, mutant M3, shows reduced activity but increased thermostability compared to wild-type enzyme	Homo sapiens
additional information	halotolerance can be generated in an enzyme solely by modifying surface residues. Rational design of carbonic anhydrase II is undertaken in three stages replacing 18 residues in total, crystal structures confirm changes are confined to surface residues. Catalytic activities and thermal unfolding temperatures of the designed enzymes increase at high salt concentrations demonstrating their shift to halotolerance, whereas the opposite response is found in the wild-type enzyme. Molecular dynamics calculations, overview	Homo sapiens
N24D/V39D/R57D/Q74E/K169E/N252D	site-directed mutagenesis, mutant M3, shows reduced activity but increased thermostability and halostability compared to wild-type enzyme	Homo sapiens
N62D	site-directed mutagenesis	Homo sapiens

General Stability

General Stability	Organism
a key role for sodium ions in increasing halotolerant enzyme stability largely through interactions with the highly ordered first Na+ hydration shell, molecular dynamics calculations, overview	Homo sapiens

Inhibitors

Inhibitors	Comment	Organism	Structure
nitrate	mutants M3 and M4 are more resistent against inhibition by nitrate compared to wild-type and mutants M1 and M2	Homo sapiens
SDS	the wild-type enzyme is inactivated at 0.4% w/v, the mutant enzymes are more stable showing residual activity at that concentration	Homo sapiens

Metals/Ions

Metals/Ions	Comment	Organism	Structure
additional information	distribution of cations and anions within 3 A of the enzyme surface, overview	Homo sapiens
Na+	a key role for sodium ions in increasing halotolerant enzyme stability largely through interactions with the highly ordered first Na+ hydration shell, molecular dynamics calculations, overview. Very strong activation by Na2SO4 of all enzymes, especially of mutant M4, slight activation by NaCl, overview	Homo sapiens
Zn2+	required, metalloenzyme	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
H2CO3	Homo sapiens	-	CO2 + H2O	-	r

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	P00918	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant wild-type and mutant enzymes from Escherichia coli	Homo sapiens

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
additional information	-	esterase activity measurements of wild-type and mutant enzymes, pH 8.5, substrate 4-nitrophenyl acetate	Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
H2CO3	-	Homo sapiens	CO2 + H2O	-	r
additional information	esterase activity assay with 4-nitrophenyl acetate as substrate for determination of activity of wild-type and mutant enzymes	Homo sapiens	?	-	?

Subunits

Subunits	Comment	Organism
More	secondary structure stability of wild-type and mutant enzymes in SDS, overview	Homo sapiens

Synonyms

Synonyms	Comment	Organism
CAII	-	Homo sapiens
carbonic anhydrase II	-	Homo sapiens
mesohalophilic carbonic anhydrase	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
42.5	-	mutant M4	Homo sapiens
50	-	mutants M2 and M3	Homo sapiens
66.5	-	mutant M1	Homo sapiens
67.5	-	wild-type enzyme	Homo sapiens

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
additional information	-	thermal stability and thermal unfolding analyses of wild-type enzyme and mutants M1-M4, overview	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8.5	-	assay at	Homo sapiens

IC50 Value

IC50 Value	IC50 Value Maximum	Comment	Organism	Inhibitor	Structure
0.01	-	pH 7.5, 20°C, inhibition of CO2 hydration reaction	Homo sapiens	nitrate

General Information

General Information	Comment	Organism
additional information	a key role for sodium ions in increasing halotolerant enzyme stability largely through interactions with the highly ordered first Na+ hydration shell, molecular dynamics calculations, overview	Homo sapiens

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Release 2023.1 (January 2023)