Enzyme carbonic anhydrase

Fig. 11.37 Free energy profile for the nucleophilic attack of water on CO2 (a) in aqueous solution and (b) in the enzyme carbonic anhydrase. (Graphs redrawn from Aqvist J, M Fothergill and A Warshel 1993. Computer Simulai of the COj/HCOf Interconversion Step in Human Carbonic Anhydrase I. Journal of the American Chemical Society 115 631-635.)...

Many important biochemical reactions involve Lewis acid Lewis base chemistry Carbon dioxide is rapidly converted to hydrogen carbonate ion m the presence of the enzyme carbonic anhydrase... 

At 20 C, the rate constant for this uncatalyzed reaction, uncat is 0.03/sec. In the presence of the enzyme carbonic anhydrase, the rate constant for this reaction, is 10 /sec. 

The enzyme carbonic anhydrase promotes the hydration of COg. Many of the protons formed upon ionization of carbonic acid are picked up by Hb as Og dissociates. The bicarbonate ions are transported with the blood back to the lungs. When Hb becomes oxygenated again in the lungs, H is released and reacts with HCO3 to re-form HgCOj, from which COg is liberated. The COg is then exhaled as a gas. 

Among one of the more unusual side effects noticed as the use of the sulfonamides became widespread was the increased urine output of many patients treated with these drugs. The fact that the urine was unusually alkaline led to the suspicion, later (Confirmed by independent means, that these agents were responsible for partial inhibition of the enzyme carbonic anhydrase. Inhibition of this enzyme causes increased excretion of sodium and bicarbonate ions as well as water, in effect bringing about diure-... 

Carbonic anhydrase is an enzyme that produces free hydrogen ions, which are then exchanged for sodium ions in the kidney tubules. Carbonic anhydrase inhibitors inhibit the action of the enzyme carbonic anhydrase This effect results in the excretion of sodium, potassium, bicarbonate, and water. Carbonic anhydrase inhibitors also decrease the production of aqueous humor in the eye, which in turn decreases intraocular pressure (IOP) (ie, the pressure within the eye). 

The approach taken above estimates the effect of the metal by simply considering its electrostatic effect (subjected, of course, to the correct steric constraint as dictated by the metal van der Waals parameters). To examine the validity of this approach for other systems let s consider the reaction of the enzyme carbonic anhydrase, whose active site is shown in Fig. 8.6. The reaction of this enzyme involves the hydration of C02, which can be described as (Ref. 5)... 

CO3 species was formed and the X-ray structure solved. It is thought that the carbonate species forms on reaction with water, which was problematic in the selected strategy, as water was produced in the formation of the dialkyl carbonates. Other problems included compound solubility and the stability of the monoalkyl carbonate complex. Van Eldik and co-workers also carried out a detailed kinetic study of the hydration of carbon dioxide and the dehydration of bicarbonate both in the presence and absence of the zinc complex of 1,5,9-triazacyclododecane (12[ane]N3). The zinc hydroxo form is shown to catalyze the hydration reaction and only the aquo complex catalyzes the dehydration of bicarbonate. Kinetic data including second order rate constants were discussed in reference to other model systems and the enzyme carbonic anhy-drase.459 The zinc complex of the tetraamine 1,4,7,10-tetraazacyclododecane (cyclen) was also studied as a catalyst for these reactions in aqueous solution and comparison of activity suggests formation of a bidentate bicarbonate intermediate inhibits the catalytic activity. Van Eldik concludes that a unidentate bicarbonate intermediate is most likely to the active species in the enzyme carbonic anhydrase.460... 

Synthesis of functional models of carbonic anhydrase has been attempted with the isolation of an initial mononuclear zinc hydroxide complex with the ligand hydrotris(3-t-butyl-5-methyl-pyrazolyl)borate. Vahrenkamp and co-workers demonstrate the functional as well as the structural analogy to the enzyme carbonic anhydrase. A reversible uptake of carbon dioxide was observed, although the unstable bicarbonate complex rapidly forms a dinuclear bridged complex. In addition, coordinated carbonate esters have been formed and hydrolyzed, and inhibition by small ions noted.462 A number of related complexes are discussed in the earlier Section 6.8.4. 

The importance of zinc may extend even further as the processes that control carbon uptake by marine phytoplankton are important in the carbon cycle, and the zinc enzyme carbonic anhydrase could be a limiting factor. The concentration of zinc available to some marine phytoplankton has... 

The carbon dioxide produced during cellular metabolism diffuses out of the cells and into the plasma. It then continues to diffuse down its concentration gradient into the red blood cells. Within these cells, the enzyme carbonic anhydrase (CA) facilitates combination of carbon dioxide and water to form carbonic acid (H2C03). The carbonic acid then dissociates into hydrogen ion (H+) and bicarbonate ion (HC03). 

Thermal reaction techniques enable a quantification of the influence of solvation on reactivities.1,2,19 One particular reaction which is a good example of how solvation can affect the nature of a core ion reaction site comes from a study118 of the interaction of OH with C02. The gas-phase reaction between the individual species is quite exothermic and can only take place by a three-body association mechanism. The reaction proceeds very slowly in the liquid phase and has been calculated119 to have a barrier of about 13 kcal mol-1. In biological systems, the reaction rate is enhanced by about 4 orders of magnitude through the enzyme carbonic anhydrase. Recent studies carried out in our laboratory provide detailed... 

L-amino acids as effident activators of zinc enzyme carbonic anhydrase. Liebigs Annalen der Chemie,... 

Part of the metabolic machinery of an osteoclast resembles the red cell and the renal tubule cells because all of these cell types contain the enzyme carbonic anhydrase (carbonate dehydratase) which generates acid, that is protons, and have ion pumps in their plasma membranes. The mechanism of bone resorption requires the action of cathepsin and metalloproteinase-9 working in an acidic environment (Figure 9.8). 

The fact that zinc is known to be a component of an enzyme carbonic anhydrase leaves no doubt as to its physiological significance. The amount of zinc in erythrocytes seems to parallel the carbonic anhydrase activity.21 The leucocytes which appear to lack carbonic anhydrase contain about 25 times as much zinc (per cell) as do the erythrocytes.20 It seems likely that an investigation of the zinc content of different types of white blood cells coupled with a study of individuals from the standpoint of the different types of white cells present (p. 35) would lead to the discovery of substantial inter-individual differences. The wide spread in the zinc concentrations in three human spleens has already been mentioned (p. 72). A recent study has been made of the intake and excretion of zinc by 13... 

A recent report (Pocker and Meany, 1964) states that the enzyme carbonic anhydrase is a powerful catalyst for the reversible hydration of acetaldehyde, but no details have yet been published. 

The Bronsted relationship can be strictly accurate only over a certain range of acid and base strengths. When has diffusion-controlled values, which of course cannot be exceeded, the linear plot of log k/ y vs log must level off to a zero slope, that is a = 0. As well as being reported, although rarely, in simple metal complexes, the resultant curvature in the Bronsted plot is also shown by the zinc enzyme carbonic anhydrase (Chap. 8. Zn(II)). In... 

Ambient concentrations of COj are very low and usually biolimiting. Hence phytoplankton generally rely on bicarbonate as their carbon source. Phytoplankton must convert this bicarbonate to COj to enable production of organic matter. This conversion is facilitated by the Zn-containing enzyme, carbonic anhydrase (Table 11.4). Some phytoplankton release carbonic anhydrase into seawater with the resulting COj then transported across their cell membrane. 

The products of chemical weathering, Ca, H4Si04, and 2HCOj, are transported by river runoff into the ocean, where they are then available to be returned to biogenic form by marine plankton. (Marine plankton have an enzyme, carbonic anhydrase, that converts bicarbonate to CO2.)... 

The first explicit treatment of the DCC concept using proteins was reported by Lehn and Hue in 1997 [1,7]. In a touchstone paper for the development of the DCC area in general, they synthesized an imine DCL to probe the active site of the enzyme carbonic anhydrase (CA). The composition of... 

One of the side effects noted in the clinical use of the sulfonamide antibacterial agents was a diuretic effect caused by inhibition of the enzyme carbonic anhydrase. Attempts to capitalize on this side effect so as to obtain agents with greatly enhanced diuretic activity first met success when a heterocyclic ring was substituted for the benzene ring of the sulfonamide. Treatment of the hydrazine derivative, 151, with phosgene leads... 

Pharmacology These agents are nonbacteriostatic sulfonamides that inhibit the enzyme carbonic anhydrase. This action reduces the rate of aqueous humor formation, resulting in decreased lOP. 

The most convincing proposal is that camosine plays one or more roles in control of intracellular hydrogen ion concentration (Abe, 2000 Vaughan-Jones et al, 2006). Camosine is an effective physiological buffer it is presumed that this property explains its predominant association with white, glycolytic, muscles which possess relatively few mitochondria and thereby generate lactic acid. Not only may camosine, also possible in its acetylated form, help to directly suppress the rise in hydrogen ion concentration but its ability to activate the enzyme carbonic anhydrase (Temperini et al, 2005) would increase bicarbonate buffer capacity. These properties may help explain camosine s protective action in ischaemia, a condition associated with severe intracellular acidosis. 

Four examples of catalytic or regulatory zinc proteins are reviewed here, and the discussion of metalloprotein function is set within the context of the metal ion and its coordination polyhedron. In the zinc enzymes carbonic anhydrase (carbonate dehydratase) II and carboxypeptidase A, the coordination polyhedron of the metal ion changes as the... 

Mechanism of Action A carbonic anhydrase inhibitor that reduces formation of hydrogen and bicarbonate ions from carbon dioxide and water by inhibiting, in proximal renal tubule, the enzyme carbonic anhydrase, thereby promoting renal excretion of sodium, potassium, bicarbonate, and water. Ocular Reduces rate of aqueous humor formation, lowers intraocular pressure. Therapeutic Effect Produces anticonvulsant activity. 

Carbonic anhydrase influences the tubular reabsorption of sodium in proximal tubule where biocarbonate absorption occurs and in the distal tubule where sodium is exchanged for potassium or hydrogen ion and bicarbonate is formed as the accompanying anion. The hydration of carbon dioxide takes place under the influence of enzyme carbonic anhydrase which forms carbonic acid which dissociates and breaks into hydrogen and carbonate ions. 

Acetazolamide inhibits the enzyme carbonic anhydrase, and interferes with the ability of the renal tubules to produce and secrete hydrogen ions. And, the diuretic action is due to the decreased sodium biocarbonate absorption in proximal tubules and diminished hydrogensodium exchange in the distal tubules. 

The most important function of the proximal tubular cell is the conservation of filtered Na+ and the reabsorption of water. The PCT is also the main site of FICO-3 reabsorption. This is achieved by the transfer of Na+ and FICO-3 from the tubular lumen into the cell and then into the extracellular fluid (ECF) accompanied by the passive reabsorption of approximately 70% of the filtered water via the tight junctions between the tubular cells. The presence of the enzyme carbonic anhydrase in the cytoplasm and luminal epithelium of the cells of the PCT allows the kidney to eliminate FI+ while simultaneously retaining FICO-3. 

The importance of the biochemistry of hydration of CO2 and dehydration of HCOg in an aqueous environment has led to extensive and invigorating research on the enzyme carbonic anhydrase pertaining to its structural details, metal ion cofactor, its coordination environment (12) and kinetic activity Model studies, both theoretical and experimental, have been undertaken using primarily the complexes of Zn(II), Mn(II), and Co(II), the latter one being its closest equivalent (13). 

It is the intention of the authors to present a brief account on metal carbonato complexes which have a direct bearing on the reversible hydration of CO2 by the enzyme carbonic anhydrase. Emphasis is placed on the integration of the kinetic and mechanistic concepts derived from the studies on model systems with the available kinetic, chemical and structural information on the enzyme carbonic anhydrase. To start, the kinetics and equilibria of dissolved CO2, relevant to the present context, are presented. 

This reaction is rather slow in the absence of the enzyme carbonic anhydrase, which is usually the case with fermentation broths, although this enzyme exists in the red blood cells. Thus, any increase of for CO, desorption from fermentation broths due to simultaneous diffusion of HCO3 seems negligible. 

Reaction (B) is very rapid. Reaction (A) is slow, but becomes very rapid in the presence ofthe enzyme carbonic anhydrase, which exists in the erythrocytes. Carbon dioxide produced by the gas exchange in tissues moves into the erythrocytes, while bicarbonate ions produced by reactions (A) and (B) in the erythrocytes move out into the plasma. 

The C02 produced by glucose oxidation is converted in erythrocytes to the more soluble HCOcT, in a reaction catalyzed by the enzyme carbonic anhydrase ... 

The protons released then react with more bicarbonate to form carbonic acid which is converted to C02 and water (Eq. 8-12) by the enzyme carbonic anhydrase. 

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