Effects of cryoprotectants on the structure and thermostability of the human carbonic anhydrase II-acetazolamide complex

Protein X‐ray crystallography has seen a progressive shift from data collection at cool/room temperature (277–298 K) to data collection at cryotemperature (100 K) because of its ease of crystal preparation and the lessening of the detrimental effects of radiation‐induced crystal damage, with 20–25%(...

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Bibliographic Details
Published in:Acta crystallographica. Section D, Biological crystallography. Biological crystallography., 2013-05, Vol.69 (5), p.860-865
Main Authors: Aggarwal, Mayank, Boone, Christopher D., Kondeti, Bhargav, Tu, Chingkuang, Silverman, David N., McKenna, Robert
Format: Article
Language:English
Subjects:
acetazolamide
Acetazolamide - chemistry
Calorimetry, Differential Scanning
Carbonic anhydrase
Carbonic Anhydrase II - antagonists & inhibitors
Carbonic Anhydrase II - chemistry
Carbonic Anhydrase II - metabolism
Carbonic Anhydrase Inhibitors - chemistry
Catalytic Domain
CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY
Cryoprotectants
Cryoprotective Agents - chemistry
Cryoprotective Agents - pharmacology
CRYSTAL STRUCTURE
CRYSTALLOGRAPHY
Crystallography, X-Ray
CRYSTALS
DAMAGE
Data acquisition
Enzyme Stability - drug effects
Enzymes
GLYCEROL
Glycerol - chemistry
Glycerol - metabolism
Glycerol - pharmacology
Humans
Inhibition
Inhibitors
KINETICS
MATHEMATICAL SOLUTIONS
Models, Molecular
Protein Conformation
PROTONS
Research Papers
SACCHAROSE
SOLUTIONS
sucrose
Sucrose - chemistry
Temperature
Thermal stability
WATER
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Summary:Protein X‐ray crystallography has seen a progressive shift from data collection at cool/room temperature (277–298 K) to data collection at cryotemperature (100 K) because of its ease of crystal preparation and the lessening of the detrimental effects of radiation‐induced crystal damage, with 20–25%(v/v) glycerol (GOL) being the preferred choice of cryoprotectant. Here, a case study of the effects of cryoprotectants on the kinetics of carbonic anhydrase II (CA II) and its inhibition by the clinically used inhibitor acetazolamide (AZM) is presented. Comparative studies of crystal structure, kinetics, inhibition and thermostability were performed on CA II and its complex with AZM in the presence of either GOL or sucrose. These results suggest that even though the cryoprotectant GOL was previously shown to be directly bound in the active site and to interact with AZM, it affects neither the thermostability of CA II nor the binding of AZM in the crystal structure or in solution. However, addition of GOL does affect the kinetics of CA II, presumably as it displaces the water proton‐transfer network in the active site.
ISSN:1399-0047
0907-4449
1399-0047
DOI:10.1107/S0907444913002771