Sem enzima catalista, a reação biológica mais lenta conhecida levaria 1 trilhão de anos!!!!

Para ler e ponderar cum granum salis a probabilidade desse evento inusitado ocorrer na natureza por mero acaso, necessidade e sem design inteligente:

The rate of hydrolysis of phosphomonoester dianions and the exceptional catalytic proficiencies of protein and inositol phosphatases

1. Chetan Lad*,
2. Nicholas H. Williams*,†, and
3. Richard Wolfenden†,‡

+Author Affiliations

1. *Centre for Chemical Biology, Krebs Institute for Biomolecular Science, Department of Chemistry, University of Sheffield, Sheffield S3 7HF, United Kingdom; and ‡Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC 27599

1.Contributed by Richard Wolfenden


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Abstract

To evaluate the proficiency of phosphatases as catalysts, the rate of the uncatalyzed hydrolysis of simple phosphate monoester dianions was estimated by extrapolating rates measured over a range of high temperatures. The rate of spontaneous hydrolysis of phenyl phosphate dianion indicates that a linear free energy relationship reported earlier is reliable for leaving groups whose conjugate acids have pKa values up to at least 10. Using Teflon reaction vessels, it proved possible to follow the hydrolysis of methyl phosphate and 3-(4-carboxy)-2,2-dimethylpropyl phosphate in strong alkali. Even in 1 M KOH, the reaction was found to be specific acid catalyzed. These results establish an upper limit for dianion reactivity, which had been overestimated earlier as a result of the leaching by alkali of silicic acid from quartz reaction vessels. The present findings indicate that the half-time for attack by water on alkyl phosphate dianions is 1.1 × 1012 years (k = 2 × 10−20 s) at 25°C and that phosphatases involved in cell signaling and regulation produce the largest rate enhancements that have been identified thus far. Protein phosphatase-1 and inositol 1-phosphatase exceed all other known enzymes in their affinities for the altered substrates in the transition state.

Most enzyme reactions proceed with k cat/K m values in the neighborhood of 107 M−1⋅s−1 and appear to be similarly efficient according to that criterion. However, to assess the proficiency of an enzyme as a catalyst, and its corresponding affinity for the altered substrate in the transition state, it is necessary to compare k cat/K m with the rate constant of the corresponding reaction under the same conditions in the absence of a catalyst. In contrast to the relatively narrow range of values of k cat/K m observed for enzyme catalyzed reaction rates, the rates of these uncatalyzed reactions span a range of at least 19 orders of magnitude (1). Thus, differences in enzyme proficiency tend to reflect differences in the rates of the uncatalyzed reactions rather than the catalyzed reactions, and enzymes differ greatly from one another in their prowess as catalysts. Enzymes that catalyze the slowest reactions are of practical interest, in that they offer sensitive targets for inhibition by transition-state analogues. Their mechanisms of action are also particularly challenging to rationalize. Conspicuous among these slow reactions is the hydrolysis of monoesters of phosphoric acid.

In biological systems, phosphate monoesters are cleaved by two groups of monoesterases that act by different mechanisms.

One group of phosphomonoesterases, typified by bacterial alkaline phosphatase (2) and protein tyrosine phosphatases (3), uses an active-site nucleophile to displace the alcohol-leaving group from the substrate to form a phosphoryl-enzyme intermediate, which is subsequently hydrolyzed. Rate enhancement by enzymes that act through a double-displacement mechanism is not simple to analyze in terms of transition-state affinity or transition-state stabilization (4), although rates of reaction with model nucleophiles can be used to estimate equilibrium constants for transition-state interchange between an enzyme and the model nucleophile (5).

A second group of phosphatases catalyzes direct attack by water on phosphorus to displace the alcohol-leaving group and therefore lends itself directly to the estimation of transition-state affinities. Enzymes that bring about direct water attack include protein phosphatase-1 of which numerous variants with differing specificities have been found within the human proteome; fructose 1,6-bisphosphatase, which catalyzes the rate-determining step in carbohydrate catabolism; and inositol 1-phosphatases, which are involved in the transmission of hormonal signals. Here, we report that these enzymes exceed all other known enzymes in their powers as catalysts and in their affinities for the altered substrate in the transition state.

In the investigation of very slow reactions, simple model substrates are helpful in avoiding side reactions that might obscure the reaction of interest. The spontaneous hydrolysis of methyl phosphate (MeP) is relatively rapid at low pH where it is present mainly as the monoanion, but proceeds more slowly with increasing pH as the abundance of the monoanion (MeP−) declines (6). In experiments in sealed quartz tubes, reported earlier (7), the rate of hydrolysis of MeP appeared to reach a constant value more than pH 10, corresponding to the reaction of the dianion (MeP−2) with water. Slow as it was, that reaction proceeded considerably more rapidly than had been expected by extrapolation of rate constants for hydrolysis of monoester dianions with much better leaving groups (Fig. 1) (8, 9). Reinvestigating that discrepancy, we have established that the spontaneous hydrolysis of MeP−2 proceeds much more slowly than was previously reported and that phosphatases are even more proficient as catalysts than had been suspected.

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Fui, cada vez mais convencido de que as atuais teorias sobre a origem e evolução do universo e da vida estão mais para especulação do que verdadeira ciência baconiana!