The crystal structure of F65A/Y131C
murine
mages/gifchars/alpha.gif" BORDER=0>-carbonic anhydrase V (CAV), covalently
modified at cysteine residues with 4-chloro
methyli
midazole, is reported at 1.88 Å resolution. This
modification introduces a
methyli
midazole (MI) group at residue C131 in the active site with i
mportantconsequences. F65A/Y131C-MI CAV exhibits an up to 3-fold enhance
ment of catalytic activity over thatof wild-type CAV [Earnhardt, J. N., Wright, S. K., Qian, M., Tu, C., Laipis, P. J., Viola, R. E., andSilver
man, D. N. (1999)
Arch. Biochem. Biophys. 361, 264-270]. In this
modified CAV variant, C131-MI acts as a proton shuttle, facilitating the deprotonation of a zinc-bound water
molecule to regeneratethe nucleophilic zinc-bound hydroxide ion. A network of three hydrogen-bonded water
molecules, acrosswhich proton transfer likely proceeds, bridges the zinc-bound water
molecule and the C131-MI i
midazolegroup. The structure of F65A/Y131C-MI CAV is co
mpared to structures of Y64H/F65A
murine CAV,wild-type hu
man
mages/gifchars/alpha.gif" BORDER=0>-carbonic anhydrase II, and the
mages/gifchars/ga
mma.gif" BORDER=0 >-carbonic anhydrase fro
m Methanosarcina thermophilain an effort to outline co
mmon features of catalytic proton shuttles.