CRYSTAL-STRUCTURE OF CARBONIC-ANHYDRASE FROM NEISSERIA-GONORRHOEAE AND ITS COMPLEX WITH THE INHIBITOR ACETAZOLAMIDE

The crystal structure of carbonic anhydrase from Neisseria gonorrhoeae has been solved to a resolution of 1.78 Angstrom by molecular replace ment using human carbonic anhydrase II as a template. After refinement the R factor was 17.8% (R-free = 23.2%). There are two molecules per asymmetric unit (space group P2(1)), but they have essentially identic al structures. The fold of the N. gonorrhoeae enzyme is very similar t o that of human isozyme II; 192 residues, 74 of which are identical in the two enzymes, have equivalent positions in the three-dimensional s tructures. This corresponds to 85% of the entire polypeptide chain of the bacterial enzyme. The only two cysteine residues in the bacterial enzyme, which has a periplasmic location in the cell, are connected by a disulfide bond. Most of the secondary structure elements present in human isozyme II are retained in N. gonorrhoeae carbonic anhydrase, b ut there are also differences, particularly in the few helical regions . Long deletions in the bacterial enzyme relative to human isozyme II have resulted in a considerable shortening of three surface loops. One of these deletions, corresponding to residues 128 to 139 in the human enzyme, leads to a widening of the entrance to the hydrophobic part o f the active site cavity. Practically all the amino acid residues in t he active site of human isozyme II are conserved in the N. gonorrhoeae enzyme and have similar structural positions. However, the imidazole ring of a histidine residue, which has been shown to function as a pro ton shuttle in the catalytic mechanism of the human enzyme, interacts with an extraneous entity, which has tentatively been identified as a 2-mercaptoethanol molecule from the crystallization medium. When this entity is removed by soaking the crystal in a different medium, the si de-chain of His66 becomes quite mobile. The structure of a complex wit h the sulfonamide inhibitor, acetazolamide, has also been determined. Its position in the active site is very similar to that observed in hu man carbonic anhydrase II. (C) 1998 Academic Press.