Spinach carbonic anhydrase: investigation of the zinc-binding ligands by site-directed mutagenesis, elemental analysis, and EXAFS

Biochemistry. 1994 Nov 8;33(44):13126-31. doi: 10.1021/bi00248a023.


The enzyme carbonic anhydrase has been well characterized in mammalian systems, but the structural properties of the plant isozymes remain elusive. To investigate the nature of the zinc-binding site in spinach carbonic anhydrase, we targeted potential zinc ligands for mutagenesis and examined the resulting enzymes for catalytic activity and stoichiometric zinc binding. In addition, we examined the wild-type protein using extended X-ray absorption fine structure analysis. Our results suggest that spinach carbonic anhydrase utilizes a Cys-His-Cys-H2O ligand scheme to bind the zinc ion at the active site.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Binding Sites
  • Carbonic Anhydrases / chemistry*
  • Carbonic Anhydrases / genetics
  • Cloning, Molecular
  • Electrophoresis, Polyacrylamide Gel
  • Ligands
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Spectrometry, X-Ray Emission
  • Spinacia oleracea / enzymology
  • Zinc / metabolism*


  • Bacterial Proteins
  • Ligands
  • Carbonic Anhydrases
  • Zinc

Associated data