[No authors listed]
The purple acid phosphatases (PAP) are binuclear metallohydrolases that catalyze the hydrolysis of a broad range of phosphomonoester substrates. The mode of substrate binding during catalysis and the identity of the nucleophile is subject to debate. Here, we used native Fe(3+)-Fe(2+) pig PAP (uteroferrin; Uf) and its Fe(3+)-Mn(2+) derivative to investigate the effect of metal ion substitution on the mechanism of catalysis. Replacement of the Fe(2+) by Mn(2+) lowers the reactivity of Uf. However, using stopped-flow measurements it could be shown that this replacement facilitates approximately a ten-fold faster reaction between both substrate and inorganic phosphate with the chromophoric Fe(3+) site. These data also indicate that in both metal forms of Uf, phenyl phosphate hydrolysis occurs faster than formation of a mu-1,3 phosphate complex. The slower rate of interaction between substrate and the Fe(3+) site relative to catalysis suggests that the substrate is hydrolyzed while coordinated only to the divalent metal ion. The likely nucleophile is a water molecule in the second coordination sphere, activated by a hydroxide terminally coordinated to Fe(3+). The faster rates of interaction with the Fe(3+) site in the Fe(3+)-Mn(2+) derivative than the native Fe(3+)-Fe(2+) form are likely mediated via a hydrogen bond network connecting the first and second coordination spheres, and illustrate how the selection of metal ions may be important in fine-tuning the function of this enzyme.
KEYWORDS: {{ getKeywords(articleDetailText.words) }}
Sample name | Organism | Experiment title | Sample type | Library instrument | Attributes | |||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
{{attr}} | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
{{ dataList.sampleTitle }} | {{ dataList.organism }} | {{ dataList.expermentTitle }} | {{ dataList.sampleType }} | {{ dataList.libraryInstrument }} | {{ showAttributeName(index,attr,dataList.attributes) }} |
{{ list.authorName }} {{ list.authorName }} |