Evolutionary optimization of the catalytic effectiveness of an enzyme

Burbaum, Jonathan J.; Raines, Ronald T.; Albery, W. John; Knowles, Jeremy R.

doi:10.1021/bi00450a009

Cited by 166 publications

(166 citation statements)

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“…Our conclusions did indeed differ from those of Brocklehurst (1977); this was because we were interpreting our results according to Burbaum et al (1989). For the acyl-enzyme mechanism [eqn.…”

contrasting

confidence: 56%

“…On the other hand, Burbaum et al (1989) infer that k+2-k+3, and that at (or near to) the diffusion limit the barrier for the second transition state is likely to be as low as it needs to be to avoid slowing the reaction, and so k l k+2. Brocklehurst (1977) did suggest that the condition k+2 > k l might be relaxed to k+2 k1 with little loss in catalytic efficiency, and so this leaves the main difference between Brocklehurst (1977) and Burbaum et al (1989) as the additional requirement that k+2=-k+3 in the latter treatment. Our experimental results for ,-lactamases (Christensen et al, 1990) were that k1 k+2-k+3, and that the reactions were partly (or largely) diffusion-controlled; thus our results were entirely consistent with the theories of Burbaum et al (1989).…”

mentioning

confidence: 99%

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Kinetic parameters of the acyl-enzyme mechanism

Waley

1990

Biochemical Journal

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contrasting

confidence: 56%

mentioning

confidence: 99%

Kinetic parameters of the acyl-enzyme mechanism

Waley

1990

Biochemical Journal

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“…Consequently, the equilibrium ratios of the bound triose phosphates dihydroxyacetone phosphate (DHAP), triose 1,2-enediol 3-phosphate (enediol), and D-glyceraldehyde 3-phosphate (GAP) should shift compared with those of free triose phosphates in aqueous solutions (14,15); i.e., the equilibrium constant of the enzymebound substrate and product will be closer to unity than the equilibrium constant of the free species. By using the rates determined by Albery and Knowles (9) for the basic kinetic model (summarized in Fig.…”

Section: Strongly Suggest That the Reaction Intermediate Is A Cis-enementioning

confidence: 99%

Substrate product equilibrium on a reversible enzyme, triosephosphate isomerase

Rozovsky

McDermott

2007

Proc. Natl. Acad. Sci. U.S.A.

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The highly efficient glycolytic enzyme, triosephosphate isomerase, is expected to differentially stabilize the proposed stable reaction species: ketone, aldehyde, and enediol(ate). The identity and steady-state populations of the chemical entities bound to triosephosphate isomerase have been probed by using solid-and solution-state NMR. The 13 C-enriched ketone substrate, dihydroxyacetone phosphate, was bound to the enzyme and characterized at steady state over a range of sample conditions. The ketone substrate was observed to be the major species over a temperature range from ؊60°C to 15°C. Thus, there is no suggestion that the enzyme preferentially stabilizes the reactive intermediate or the product. The predominance of dihydroxyacetone phosphate on the enzyme would support a mechanism in which the initial proton abstraction in the reaction from dihydroxyacetone phosphate to D-glyceraldehyde 3-phosphate is significantly slower than the subsequent chemical steps.enzymatic catalysis ͉ Michaelis complex ͉ solid-state NMR T he glycolytic enzyme triosephosphate isomerase (TIM) catalyzes isomerization of a ketone to an aldehyde, progressing via two successive proton transfers involving carbon acids and additional proton transfers for the attached oxygen atoms (Fig. 1). The precise path of the transferred proton and the identity of the reaction intermediates have been, by and large, deduced from the fate of isotopic labels (1, 2). The existence of a stable enediol(ate) intermediate has been inferred from the loss of an isotope label from the substrate during the reaction (1, 3); this reaction intermediate must exist with a sufficient lifetime to allow partial equilibration of the catalytic base with the surrounding solvent. Moreover, when the reaction takes place in tritiated water the solvent's tritium is incorporated into both the substrate and product, evidence that the hydrogen of the newly formed carbonhydrogen bond is mostly derived from the solvent (4, 5) and that exchange occurs from an enzyme bound intermediate. The stereochemistry of the reaction (2) and the orientation of the substrate in the Michaelis complex (6) strongly suggest that the reaction intermediate is a cis-enediol(ate). Based on a comprehensive study of isotope effects supplemented by extensive biophysical investigations of the reaction mechanism (7, 8), Albery and Knowles (9) constructed a kinetically detailed energy profile for the reversible interconversion of substrate, product, and one intermediate subject to isotope exchange with solvent ( Fig. 1).This now well known reaction energy profile illustrated for the first time many of the essential elements contributing to the high catalytic efficiency brought about by enzymes (10). For example, the free-energy profile illustrates the potential catalytic power of preferential stabilization of intermediates in a reaction. The high catalytic efficiency of TIM [almost 10 orders of magnitude compared with the reaction catalyzed by an acetate ion (11)] is thought to arise from matching the pK a va...

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“…and the nature, solubility or availability of substrates. It is also quite possible that our concept of catalytic effectiveness remains imperfect and may require more sophisticated models as those developed by Knowles and coworkers [47]. The peculiar case of the class C i-lactamase (cephalosporinase) from Psychrobacter immobilis also deserves some comments because the specific activity of this antibiotic-degrading enzyme at low temperatures is not favoured when compared to i-lactamases from mesophilic pathogenic bacteria ( fig.…”

Section: Kinetic and Thermodynamic Optimizationmentioning

confidence: 99%

Psychrophilic enzymes: molecular basis of cold adaptation

Feller

Gerday

1997

CMLS, Cell. mol. life sci.

371

233

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Abstract. Psychrophilic organisms have successfully provides enhanced abilities to undergo conformational colonized polar and alpine regions and are able to grow changes during catalysis. Thermal instability of coldefficiently at sub-zero temperatures. At the enzymatic adapted enzymes is therefore regarded as a consequence level, such organisms have to cope with the reduction of of their conformational flexibility. A survey of the psychrophilic enzymes studied so far reveals only minor chemical reaction rates induced by low temperatures in alterations of the primary structure when compared to order to maintain adequate metabolic fluxes. Thermal compensation in cold-adapted enzymes is reached mesophilic or thermophilic homologues. However, all known structural factors and weak interactions inthrough improved turnover number and catalytic effivolved in protein stability are either reduced in number ciency. This optimization of the catalytic parameters can originate from a highly flexible structure which or modified in order to increase their flexibility.Key words. Psychrophiles; extremophiles; cold adaptation; microbial proteins; protein stability; weak interactions; Antarctic.Psychrophiles live at the lowest temperatures which allow the development of living organisms. These extremophilic organisms, either prokaryotic or eukaryotic, represent a major class of the living world if we consider the vast extent of permanently cold environments on Earth, such as polar and alpine regions or deep-sea waters. The successful colonization of such severe ecological niches by psychrophiles, their diversity and abundance are now well recognized. The lower temperature limit of life is commonly defined as the freezing point of cellular water. Although apparently simple by definition, this limit can drop significantly below 0°C, the freezing point of pure water. For instance, the freezing point of a typical marine teleost ranges between −0.5 and −0.9°C. Sodium chloride is responsible for about 85% of the freezing point depression, and the remaining 15% is due to other small solutes. Synthesis of antifreeze glycoproteins and peptides can further depress the freezing point of body fluids or cellular water. These proteins lower the freezing point by a noncolligative mechanism without altering the melting point significantly, and are therefore also referred to as thermal hysteresis proteins [1,2]. They were first discovered in the blood sera of Antarctic fish living perennially at about −1.9°C, but have been now reported in many organisms including plants, insects, fungi and bacteria [3][4][5]. Levels of thermal hysteresis activity generally range from 0.1 to 0.3°C in bacteria, from 0.2 to 0.5°C in plants, from 0.7 to 1.5°C in fish and from 3 to 6°C in insects. Besides the synthesis of cryoprotectants which lead to freeze avoidance, several organisms have developed mechanisms of freeze tolerance, involving drastic metabolic modifica-* Corresponding author.

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Evolutionary optimization of the catalytic effectiveness of an enzyme

Cited by 166 publications

References 25 publications

Kinetic parameters of the acyl-enzyme mechanism

Kinetic parameters of the acyl-enzyme mechanism

Substrate product equilibrium on a reversible enzyme, triosephosphate isomerase

Psychrophilic enzymes: molecular basis of cold adaptation

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