3RLD

Crystal structure of the Y7I mutant of human carbonic anhydrase II

Summary for 3RLD

• Entry DOI: 10.2210/pdb3rld/pdb
• Related: 2NXT
• Descriptor: Carbonic anhydrase 2, ZINC ION, SULFATE ION, ... (4 entities in total)
• Functional Keywords: zinc metalloenzyme, histidine ligands, y7i mutation, lyase
• Biological source: Homo sapiens (human)
• Cellular location: Cytoplasm : P00918
• Total number of polymer chains: 1
• Total formula weight: 29592.64

Authors

Avvaru, B.S.,Mikulski, R.,McKenna, R. (deposition date: 2011-04-19, release date: 2012-02-08, Last modification date: 2023-09-13)

Primary citation

Mikulski, R.,Avvaru, B.S.,Tu, C.,Case, N.,McKenna, R.,Silverman, D.N.
Kinetic and crystallographic studies of the role of tyrosine 7 in the active site of human carbonic anhydrase II.
Arch.Biochem.Biophys., 506:181-187, 2011

PubMed Abstract: The rate limiting step in catalysis of bicarbonate dehydration by human carbonic anhydrase II (HCA II) is an intramolecular proton transfer from His64 to the zinc-bound hydroxide. We have examined the role of Tyr7 using site-specific mutagenesis and measuring catalysis by the ¹⁸O exchange method using membrane inlet mass spectrometry. The side chain of Tyr7 in HCA II extends into the active-site cavity about 7 Å from the catalytic zinc atom. Replacement of Tyr7 with eight other amino acids had no effect on the interconversion of bicarbonate and CO₂, but in some cases caused enhancements in the rate constant of proton transfer by nearly 10-fold. The variant Y7I HCA II enhanced intramolecular proton transfer approximately twofold; its structure was determined by X-ray crystallography at 1.5 Å resolution. No changes were observed in the ordered solvent structure in the active-site cavity or in the conformation of the side chain of the proton shuttle His64. However, the first 11 residues of the amino-terminal chain in Y7I HCA II assumed an alternate conformation compared with the wild type. Differential scanning calorimetry showed variants at position 7 had a melting temperature approximately 8 °C lower than that of the wild type.

PubMed: 21145876
DOI: 10.1016/j.abb.2010.12.004

Experimental method

X-RAY DIFFRACTION (1.5 Å)