PMID- 10413479
OWN - NLM
STAT- MEDLINE
DCOM- 19990802
LR  - 20160603
IS  - 0006-2960 (Print)
IS  - 0006-2960 (Linking)
VI  - 38
IP  - 28
DP  - 1999 Jul 13
TI  - Metal binding specificity in carbonic anhydrase is influenced by conserved 
      hydrophobic core residues.
PG  - 9054-62
AB  - The role of highly conserved aromatic residues surrounding the zinc binding site 
      of human carbonic anhydrase II (CAII) in determining the metal ion binding 
      specificity of this enzyme has been examined by mutagenesis. Residues F93, F95, 
      and W97 are located along a beta-strand containing two residues that coordinate 
      zinc, H94 and H96, and these aromatic amino acids contribute to the high zinc 
      affinity and slow zinc dissociation rate constant of CAII [Hunt, J. A., and 
      Fierke, C. A. (1997) J. Biol. Chem. 272, 20364-20372]. Substitutions of these 
      aromatic amino acids with smaller side chains enhance the copper affinity (up to 
      100-fold) while decreasing the affinity of both cobalt and zinc, thereby altering 
      the metal binding specificity up to 10(4)-fold. Furthermore, the free energy of 
      the stability of native CAII, determined by solvent-induced denaturation, 
      correlates positively with increased hydrophobicity of the amino acids at 
      positions 93, 95, and 97 as well as with cobalt and zinc affinity. Conversely, 
      increased copper affinity correlates with decreased protein stability. Zinc 
      specificity is therefore enhanced by formation of the native enzyme structure. 
      These data suggest that the hydrophobic cluster in CAII is important for 
      orienting the histidine residues to stabilize metals bound with a distorted 
      tetrahedral geometry and to destabilize the trigonal bipyramidal geometry of 
      bound copper. Knowledge of the structural factors that lead to high metal ion 
      specificity will aid in the design of metal ion biosensors and de novo catalytic 
      sites.
FAU - Hunt, J A
AU  - Hunt JA
AD  - Department of Biochemistry, Duke University Medical Center, Durham, North 
      Carolina 27710, USA.
FAU - Ahmed, M
AU  - Ahmed M
FAU - Fierke, C A
AU  - Fierke CA
LA  - eng
GR  - GM17467/GM/NIGMS NIH HHS/United States
GR  - GM40602/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Biochemistry
JT  - Biochemistry
JID - 0370623
RN  - 0 (Amino Acids)
RN  - 0 (Apoenzymes)
RN  - 0 (Metalloproteins)
RN  - 3G0H8C9362 (Cobalt)
RN  - 47E5O17Y3R (Phenylalanine)
RN  - 4QD397987E (Histidine)
RN  - 789U1901C5 (Copper)
RN  - 8DUH1N11BX (Tryptophan)
RN  - EC 4.2.1.1 (Carbonic Anhydrases)
RN  - J41CSQ7QDS (Zinc)
SB  - IM
MH  - Amino Acids/*chemistry/genetics/*metabolism
MH  - Apoenzymes/chemistry/metabolism
MH  - Binding Sites
MH  - Carbonic Anhydrases/*chemistry/genetics/*metabolism
MH  - Cobalt/metabolism
MH  - Conserved Sequence
MH  - Copper/metabolism
MH  - Enzyme Stability/genetics
MH  - Histidine/metabolism
MH  - Humans
MH  - Kinetics
MH  - Metalloproteins/chemistry/genetics/metabolism
MH  - Models, Molecular
MH  - Mutagenesis, Insertional
MH  - Phenylalanine/metabolism
MH  - Protein Denaturation
MH  - Spectrum Analysis
MH  - Substrate Specificity/genetics
MH  - Tryptophan/metabolism
MH  - Zinc/*chemistry/*metabolism
EDAT- 1999/07/22 00:00
MHDA- 1999/07/22 00:01
CRDT- 1999/07/22 00:00
PHST- 1999/07/22 00:00 [pubmed]
PHST- 1999/07/22 00:01 [medline]
PHST- 1999/07/22 00:00 [entrez]
AID - bi9900166 [pii]
AID - 10.1021/bi9900166 [doi]
PST - ppublish
SO  - Biochemistry. 1999 Jul 13;38(28):9054-62. doi: 10.1021/bi9900166.