Conformational mobility of His-64 in the Thr-200 yields Ser mutant of human carbonic anhydrase II
- Duke Univ., Durham, NC (United States)
- Univ. of Pennsylvania, Philadelphia (United States)
The three-dimensional structure of the Thr-200{yields} Ser (T200S) mutant of human carbonic anhydrase II (CAII) has been determined by X-ray crystallographic methods at 2.1-{angstrom} resolution. This particular mutant of CAII exhibits CO{sub 2} hydrase activity that is comparable to that of the wild-type enzyme with a 2-fold stabilization of the E{center dot}HCO{sub 3}{minus} complex and esterase activity that is 4-fold greater than that of the wild-type enzyme. The structure of the mutant enzyme reveals no significant local changes accompanying the conservative T200S substitution, but an important nonlocal structural change is evident: the side chain of catalytic residue his-64 rotates away from the active site by 105{degree} about {chi}{sub 1} and apparently displaces a water molecule. The displaced water molecule is present in the wild-type enzyme; however, the electron density into which this water is built is interpretable as an alternate conformation of His-64 with 10-20% occupancy. The rate constants for proton transfer from the zinc-water ligand to His-64 and from His-64 to bulk solvent are maintained in the T200S variant; therefore, if His-64 is conformationally mobile about {chi}{sub 1} and/or {chi}{sub 2} during catalysis compensatory changes in solvent configuration must sustain efficient proton transfer.
- OSTI ID:
- 5602319
- Journal Information:
- Biochemistry; (United States), Vol. 30:38; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
CARBONIC ANHYDRASE
GENE MUTATIONS
ESTERASES
ENZYME ACTIVITY
CONFORMATIONAL CHANGES
CRYSTALLOGRAPHY
ELECTRON DENSITY
HISTIDINE
LIGANDS
MAN
MOLECULAR STRUCTURE
SERINE
THREONINE
X-RAY DIFFRACTION
AMINO ACIDS
ANIMALS
AZOLES
CARBON-OXYGEN LYASES
CARBOXYLIC ACIDS
COHERENT SCATTERING
DIFFRACTION
ENZYMES
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
HYDRO-LYASES
HYDROLASES
HYDROXY ACIDS
IMIDAZOLES
LYASES
MAMMALS
MUTATIONS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PRIMATES
PROTEINS
SCATTERING
VERTEBRATES
550400* - Genetics