Literature summary for 4.2.1.1 extracted from

Koruza, K.; Lafumat, B.; Vegvari, A.; Knecht, W.; Fisher, S.Z.
Deuteration of human carbonic anhydrase for neutron crystallography Cell culture media, protein thermostability, and crystallization behavior (2018), Arch. Biochem. Biophys., 645, 26-33 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene CA2, recombinant expression in Escherichia coli strain BL21(DE3), optimization of expression media and cell culture conditions to produce high levels of 3 different deuterated human carbonic anhydrases (hCAs). The labeled hCAs are then characterized and tested for deuterium incorporation by mass spectrometry, temperature stability, and propensity to crystallize, overview	Homo sapiens
gene CA9, recombinant expression in Escherichia coli strain BL21(DE3), optimization of expression media and cell culture conditions to produce high levels of 3 different deuterated human carbonic anhydrases (hCAs). The labeled hCAs are then characterized and tested for deuterium incorporation by mass spectrometry, temperature stability, and propensity to crystallize, overview. Deuteration highly reduces the enzyme yield	Homo sapiens

Cloned (Comment)

Organism

gene CA2, recombinant expression in Escherichia coli strain BL21(DE3), optimization of expression media and cell culture conditions to produce high levels of 3 different deuterated human carbonic anhydrases (hCAs). The labeled hCAs are then characterized and tested for deuterium incorporation by mass spectrometry, temperature stability, and propensity to crystallize, overview

Homo sapiens

gene CA9, recombinant expression in Escherichia coli strain BL21(DE3), optimization of expression media and cell culture conditions to produce high levels of 3 different deuterated human carbonic anhydrases (hCAs). The labeled hCAs are then characterized and tested for deuterium incorporation by mass spectrometry, temperature stability, and propensity to crystallize, overview. Deuteration highly reduces the enzyme yield

Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant differently isotope-labeled isozyme hCAII, 14 mg/ml protein, both vapour diffusion and microbatch under-oil techniques are valuated, hanging and sitting drop set-ups with 0.01 ml drops, microseeding, deuteration of hCA II does not affect the crystal structure. Large, single crystals grow for both H and D enzyme versions in microbatch under-oil experiments at pH 8.5, room temperature, X-ray diffraction structure determination and analysis	Homo sapiens
purified recombinant differently isotope-labeled isozyme hCAIX, 17 mg/ml protein, both vapour diffusion and microbatch under-oil techniques are valuated, hanging and sitting drop set-ups with 0.01 ml drops, microseeding, room temperature, deuteration of hCA XI does not affect the crystal structure, but for D-hCA IX mimic no crystals appear in any of the under-oil batch trays at all pHs tested, while H-hCA IX SV gives several good crystals per drop at pH 8.5. X-ray diffraction structure determination and analysis	Homo sapiens

Crystallization (Comment)

Organism

purified recombinant differently isotope-labeled isozyme hCAII, 14 mg/ml protein, both vapour diffusion and microbatch under-oil techniques are valuated, hanging and sitting drop set-ups with 0.01 ml drops, microseeding, deuteration of hCA II does not affect the crystal structure. Large, single crystals grow for both H and D enzyme versions in microbatch under-oil experiments at pH 8.5, room temperature, X-ray diffraction structure determination and analysis

Homo sapiens

purified recombinant differently isotope-labeled isozyme hCAIX, 17 mg/ml protein, both vapour diffusion and microbatch under-oil techniques are valuated, hanging and sitting drop set-ups with 0.01 ml drops, microseeding, room temperature, deuteration of hCA XI does not affect the crystal structure, but for D-hCA IX mimic no crystals appear in any of the under-oil batch trays at all pHs tested, while H-hCA IX SV gives several good crystals per drop at pH 8.5. X-ray diffraction structure determination and analysis

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
additional information	optimization of expression media and cell culture conditions to produce high levels of 3 different deuterated human carbonic anhydrases (hCAs). The labeled hCAs are then characterized and tested for deuterium incorporation by mass spectrometry, temperature stability, and propensity to crystallize, overview. Deuteration highly reduces the enzyme yield	Homo sapiens
additional information	optimization of expression media and cell culture conditions to produce high levels of 3 different deuterated human carbonic anhydrases (hCAs). The labeled hCAs are then characterized and tested for deuterium incorporation by mass spectrometry, temperature stability, and propensity to crystallize, overview. The protein appears to be more temperature stable at pH 7.5-8.5 compared to pH 5.5	Homo sapiens

Inhibitors

Inhibitors	Comment	Organism	Structure
additional information	presence of D2O reduces catalysis, but does not change how it proceeds; presence of D2O reduces catalysis, but does not change how it proceeds	Homo sapiens

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	steady-state kinetics and isotope effects of recombinant enzyme variants, overview	Homo sapiens

Metals/Ions

Metals/Ions	Comment	Organism	Structure
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	-	-
Homo sapiens	Q16790	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant enzyme from Escherichia coli strain BL21(DE3) by p-amino-methyl-benzenesulfonamide affinity chromatography and gel filtration, followed by ultrafiltration	Homo sapiens

Source Tissue

Source Tissue	Comment	Organism	Textmining
solid cancer cell	isoform hCA IX is not usually expressed in healthy tissues but is found in over 30 kinds of solid tumor cancers to date	Homo sapiens	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
H2CO3	-	Homo sapiens	CO2 + H2O	-	r
additional information	determination of enzyme esterase activity using p-nitrophenyl acetate as substrate. Presence of D2O reduces catalysis, but does not change how it proceeds	Homo sapiens	?	-	?

Synonyms

Synonyms	Comment	Organism
CA2	-	Homo sapiens
CA9	-	Homo sapiens
carbonic anhydrase 2	-	Homo sapiens
carbonic anhydrase 9	-	Homo sapiens
HCA II	-	Homo sapiens
hCA IX	-	Homo sapiens

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
40	60	thermal unfolding of recombinant isotope-labeled isozyme at different pH values of pH 5.5-8.5, overview	Homo sapiens
40	60	thermal unfolding of recombinant isotope-labeled isozyme at different pH values of pH 5.5-8.5, overview. The protein appears to be more temperature stable at pH 7.5-8.5 compared to pH 5.5	Homo sapiens