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 (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 | 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 | 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 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 | Comment | Organism | Structure |
---|---|---|---|
Zn2+ | required, metalloenzyme | Homo sapiens |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
H2CO3 | Homo sapiens | - |
CO2 + H2O | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | P00918 | - |
- |
Homo sapiens | Q16790 | - |
- |
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 | 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 | 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 | 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 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 |