1ELZ: E. Coli Alkaline Phosphatase Mutant (S102g)

Escherichia coli alkaline phosphatase (EC is a non-specific phosphomonoesterase that catalyzes the hydrolysis reaction via a phosphoseryl intermediate to produce inorganic phosphate and the corresponding alcohol. We investigated the nature of the primary nucleophile, fulfilled by the deprotonated Ser102, in the catalytic mechanism by mutating this residue to glycine, alanine and cysteine. The efficiencies of the S102G, S102A and S102C enzymes were 6 x 10(5)-fold, 10(5)-fold and 10(4)-fold lower than the wild-type enzyme, respectively, as measured by the kcat/Km ratio, still substantially higher than the non-catalyzed reaction. In order to investigate the structural details of the altered active site, the enzymes were crystallized and their structures determined. The enzymes crystallized in a new crystal form corresponding to the space group P6322. Each structure has phosphate at each active site and shows little departure from the wild-type model. For the S102G and S102A enzymes, the phosphate occupies the same position as in the wild-type enzyme, while in the S102C enzyme it is displaced by 2.5 A. This kinetic and structural study suggests an explanation for differences in catalytic efficiency of the mutant enzymes and provides a means to study the nature and strength of different nucleophiles in the same environment. The analysis of these results provides insight into the mechanisms of other classes of phosphatases that do not utilize a serine nucleophile.
PDB ID: 1ELZDownload
MMDB ID: 7681
PDB Deposition Date: 1998/2/10
Updated in MMDB: 2011/11 
Experimental Method:
x-ray diffraction
Resolution: 2.8  Å
Source Organism:
Similar Structures:
Biological Unit for 1ELZ: dimeric; determined by author and by software (PISA)
Molecular Components in 1ELZ
Label Count Molecule
Proteins (2 molecules)
Alkaline Phosphatase(Gene symbol: phoA)
Molecule annotation
Chemicals (8 molecules)
* Click molecule labels to explore molecular sequence information.

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