Thermodynamics of reactions catalyzed by anthranilate synthase.

Microcalorimetry and high performance liquid chromatography have been used to conduct a thermodynamic investigation of reactions catalyzed by anthranilate synthase, the enzyme located at the first step in the biosynthetic pathway leading from chorismate to tryptophan. One of the overall biochemical reactions catalyzed by anthranilate synthase is: chorismate(aq) + ammonia(aq) = anthranilate(aq) + pyruvate(aq) + H2O(l). This reaction can be divided into two partial reactions involving the intermediate 2-amino-4-deoxyisochorismate (ADIC): chorismate(aq) + ammonia(aq) = ADIC(aq) + H2O(l) and ADIC(aq) = anthranilate(aq) + pyruvate(aq). The native anthranilate synthase and a mutant form of it that is deficient in ADIC lyase activity but has ADIC synthase activity were used to study the overall ammonia-dependent reaction and the first of the above two partial reactions, respectively. Microcalorimetric measurements were performed on the overall reaction at a temperature of 298.15 K and pH 7.79. Equilibrium measurements were performed on the first partial (ADIC synthase) reaction at temperatures ranging from 288.15 to 302.65 K, and at pH values from 7.76 to 8.08. The results of the equilibrium and calorimetric measurements were analyzed in terms of a chemical equilibrium model that accounts for the multiplicity of ionic states of the reactants and products. These calculations gave thermodynamic quantities at the temperature 298.15 K and an ionic strength of zero for chemical reference reactions involving specific ionic forms. For the reaction: chorismate2-(aq) + NH4+(aq) = anthranilate-(aq) + pyruvate-(aq) + H+(aq) + H2O(l), delta rHmo = -(116.3 +/- 5.4) kJ mol-1. For the reaction: chorismate2-(aq) + NH4+(aq) = ADIC-(aq) + H2O(l), K = (20.3 +/- 4.5) and delta rHmo = (7.5 +/- 0.6) kJ mol-1. Thermodynamic cycle calculations were used to calculate thermodynamic quantities for three additional reactions that are pertinent to this branch point of the chorismate pathway. The quantities obtained in this study permit the calculation of the position of equilibrium of these reactions as a function of temperature, pH, and ionic strength. Values of the apparent equilibrium constants and the standard transformed Gibbs energy changes delta rG'mo under approximately physiological conditions are given.

A thermodynamic investigation of reactions catalyzed by tryptophan synthase.

Microcalorimetry and high-performance liquid chromatography have been used to conduct a thermodynamic investigation of the following reactions catalyzed by the tryptophan synthase alpha 2 beta 2 complex (EC 4.2.1.20) and its subunits: indole(aq) + L-serine(aq) = L-tryptophan(aq) + H2O(1); L-serine(aq) = pyruvate(aq) + ammonia(aq); indole(aq) + D-glyceraldehyde 3-phosphate(aq) = 1-(indol-3-yl)glycerol 3-phosphate(aq); L-serine(aq) + 1-(indol-3-yl)glycerol 3-phosphate(aq) = L-tryptophan(aq) + D-glyceraldehyde 3-phosphate(aq) + H2O(1). The calorimetric measurements led to standard molar enthalpy changes for all four of these reactions. Direct measurements yielded an apparent equilibrium constant for the third reaction; equilibrium constants for the remaining three reactions were obtained by using thermochemical cycle calculations. The results of the calorimetric and equilibrium measurements were analyzed in terms of a chemical equilibrium model that accounted for the multiplicity of the ionic states of the reactants and products. Thermodynamic quantities for chemical reference reactions involving specific ionic forms have been obtained. These quantities permit the calculation of the position of equilibrium of the above four reactions as a function of temperature, pH, and ionic strength. Values of the apparent equilibrium constants and standard transformed Gibbs free energy changes delta r G'(m) degree under approximately physiological conditions are given. Le Châtelier's principle provides an explanation as to why, in the metabolic pathway leading to the synthesis of L-tryptophan, the third reaction proceeds in the direction of formation of indole and D-glyceraldehyde 3-phosphate even though the apparent equilibrium constant greatly favors the formation of 1-(indol-3-yl)glycerol 3-phosphate.

Thermodynamics of the hydrolysis and cyclization reactions of alpha-, beta-, and gamma-cyclodextrin.

A thermodynamic investigation of the hydrolysis and cyclization reactions of cyclomaltohexa-, hepta-, and octa-ose (alpha-, beta-, and gamma-cyclodextrins) has been performed using microcalorimetry and high-performance liquid-chromatography. The calorimetric measurements lead to standard molar enthalpy changes delta rHm0 (T = 298.15 K, KH2PO4 buffer (m = 0.10 mol kg-1), pH = 4.58 to 5.15) for the following reactions: alpha-cyclodextrin(aq) + 6H2O(l) = 6 D-glucose(aq), beta-cyclodextrin(aq) + 7H2O(l) = 7 D-glucose(aq), gamma-cyclodextrin(aq) + 8H2O(l) = 8 D-glucose(aq). Equilibrium constants were determined for the following generalized cyclization reactions (T = 329.6 K, 0.005 mol kg-1 K2HPO4 buffer adjusted to pH = 5.55 with H3PO4) catalyzed by cyclomaltodextrin glucanotransferase: Gu(aq) = alpha-cyclodextrin(aq) + G(u-6)(aq), Gv(aq) = beta-cyclodextrin(aq) + G(v-7)(aq), Gw(aq) = gamma-cyclodextrin(aq) + G(w-8)(aq). Here, G1 is D-glucose and the Gn's (n is a positive integer) are linear maltodextrins; u, v, and w are, respectively, integers > or = 7, > or = 8, and > or = 9. Values of the equilibrium constants, standard molar Gibbs energy change delta rGm0, standard molar enthalpy change delta rHm0, standard molar entropy change delta rSm0, and standard molar heat-capacity change delta rCp,m0 are tabulated for the above reactions at T = 298.15 K. The values of delta rGm0 and delta rSm0 for the first three above-mentioned reactions rely upon an estimated value of delta rSm0 for the hydrolysis reaction of maltose to D-glucose. The thermodynamics of the disproportionation reaction Gm(aq) + Gn(aq) = Gm-1(aq) + Gn+1(aq) is also discussed. Values of the quantities delta rHm0/N, delta rGm0/N, delta rSm0/N, and delta rCp,m0/N for the three above-mentioned hydrolysis reactions where N is the number of (1-->4)-alpha-D-glucosidic bonds broken in each of these reactions, have been calculated and compared with thermodynamic quantities for the similar hydrolysis reaction of a linear oligosaccharide.

Tetralin as a substrate for camphor (cytochrome P450) 5-monooxygenase.

Camphor (cytochrome P450) 5-monooxygenase, originally isolated from the bacterium Pseudomonas putida PgG 786, catalyzes the essentially stereospecific conversion of tetralin (1,2,3,4-tetrahydronaphthalene) to (R)-1-tetralol ((R).(-)-1,2,3,4-tetrahydro-1-naphthol): tetralin(aq) + NADH(aq) + O2(aq) = (R)-1-tetralol(aq) + NAD(aq) + H2O(l). The ratio of the amount of (S)-1-tetralol to the amount of (R)-1-tetralol is small (approximately 0.04) and the reaction is essentially stereospecific. The reaction time-course plot indicates the formation of additional product(s) from the (R)-1-tetralol. It is found that the above reaction obeys Michaelis-Menten kinetics and that dimethyl sulfoxide, methanol, and p-dioxane serve as accelerators. Approximate values of a Michaelis constant Km, limiting rate Vmax, and catalytic constant kcat are obtained for this reaction under a specified set of conditions. It is shown by means of a thermochemical cycle calculation that the apparent equilibrium constant for this reaction is approximately 4 x 10(65) at T = 298.15 K and pH 7.3. Thus, this reaction is "irreversible" and, unless the enzyme system is inactivated, it will proceed in the direction of complete formation of 1-tetralol from tetralin. A detailed description of the preparation of the camphor (cytochrome P450) 5-monooxygenase enzyme system from recombinant microorganisms is given.

Thermodynamics of the hydrolysis of penicillin G and ampicillin.

Apparent equilibrium constants and calorimetric enthalpies of reaction have been measured for the beta-lactamase catalyzed hydrolysis of penicillin G(aq) and ampicillin(aq) to penicillinoic acid(aq) and to ampicillinoic acid(aq), respectively. High-pressure liquid-chromatography and microcalorimetry were used to perform these measurements. The results for the reference reactions at T = 298.15 K and Im = 0 are: Ko = (9.4 +/- 3.1) x 10(-7), delta rGo = (34.4 +/- 1.0)kJ mol-1, delta rHo = -(73.7 +/- 0.4)kJ mol-1, and delta rSo = -(363 +/- 4) J K-1 mol-1 for penicillin G-(aq) + H2O(1) = penicillinoic acid2-(aq) + H+(aq); Ko = (6.0 +/- 3.0) x 10(-6), delta rGo = (29.8 +/- 1.7) kJ mol-1, delta rHo = -(70.0 +/- 7.5) kJ mol-1, and delta rSo = -(335 +/- 26) J K-1 mol-1 for ampicillin-(aq)+ H2O(1) = ampicillinoic acid2-(aq)+H+(aq). Calorimetric enthalpies of reaction for the beta-lactamase catalyzed hydrolysis of cephalosporin C have also been measured but the reaction products have not been identified and the measured enthalpies cannot be assigned to a specific reaction. Acidity constants for ampicillin, penicillin G, ampicillinoic acid, and penicillinoic acid are also reported. A strain energy of 116 kJ mol-1 for the beta-lactam ring is obtained from thermochemical data.

Thermodynamics of the disproportionation of adenosine 5'-diphosphate to adenosine 5'-triphosphate and adenosine 5'-monophosphate. I. Equilibrium model.

The thermodynamic treatment of the disproportionation reaction of adenosine 5'-diphosphate to adenosine 5'-triphosphate and adenosine 5'-monophosphate is discussed in terms of an equilibrium model which includes the effects of the multiplicity of ionic and metal bound species and the presence of long range electrostatic and short range repulsive interactions. Calculated quantities include equilibrium constants, enthalpies, heat capacities, entropies, and the stoichiometry of the overall reaction. The matter of how these calculations can be made self-consistent with respect to both calculated values of the ionic strength and the molality of the free magnesium ion is discussed. The thermodynamic data involving proton and magnesium-ion binding data for the nucleotides involved in this reaction have been evaluated.

Thermodynamics of the disproportionation of adenosine 5'-diphosphate to adenosine 5'-triphosphate and adenosine 5'-monophosphate, II. Experimental data.

High-pressure liquid-chromatography and microcalorimetry have been used to determine equilibrium constants and enthalpies of reaction for the disproportionation reaction of adenosine 5'-diphosphate (ADP) to adenosine 5'-triphosphate (ATP) and adenosine 5'-monophosphate (AMP). Adenylate kinase was used to catalyze this reaction. The measurements were carried out over the temperature range 286 to 311 K, at ionic strengths varying from 0.06 to 0.33 mol kg(-1), over the pH range 6.04 to 8.87, and over the pMg range 2.22 to 7.16, where pMg = -log a(Mg2+). The equilibrium model developed by Goldberg and Tewari (see the previous paper in this issue) was used for the analysis of the measurements. Thus, for the reference reaction: 2 ADp(3-) (ao) AMp(2-) (ao)+ ATp- (ao), K degrees = 0.225 +/- 0.010, DeltaG degrees = 3.70 +- 0.11 kJ mol (-1), DeltaH degrees = -1.5 +/- 1. 5 kJ mol (-1), degrees S degrees = -17 +/- 5 J mol(-1)K(-1), and ACP(p) degrees approximately = -46 J mo1l(-1)K(-1) at 298.15 K and 0.1 MPa. These results and the thermodynamic parameters for the auxiliary equilibria in solution have been used to model the thermodynamics of the disproportionation reaction over a wide range of temperature, pH, ionic strength, and magnesium ion morality. Under approximately physiological conditions (311.15 K, pH 6.94, [Mg2+] = 1.35 x 10(-3) mol kg(-1), and I = 0.23 mol kg(-1)) the apparent equilibrium constant (KA' = m(SigmaAMP)m(SigmaATP)/[ m(SigmaADP)]2) for the overall disproportionation reaction is equal to 0.93 +/- 0.02. Thermodynamic data on the disproportionation reaction and literature values for this apparent equilibrium constant in human red blood cells are used to calculate a morality of 1.94 x 10(-4) mol kg(-1) for free magnesium ion in human red blood cells. The results are also discussed in relation to thermochemical cycles and compared with data on the hydrolysis of the guanosine phosphates.

Thermodynamics of hydrolysis of disaccharides. Lactulose, alpha-D-melibiose, palatinose, D-trehalose, D-turanose and 3-o-beta-D-galactopyranosyl-D-arabinose.

High-pressure liquid chromatography and microcalorimetry have been used to study the thermodynamics of the hydrolysis reactions of a series of disaccharides. The enzymes used to bring about the hydrolyses were: beta-galactosidase for lactulose and 3-o-beta-D-galactopyranosyl-D-arabinose; beta-glucosidase for alpha-D-melibiose; beta-amylase for D-trehalose; isomaltase for palatinose; and alpha-glucosidase for D-turanose. The buffer used was sodium acetate (0.02-0.10 M and pH 4.44-5.65). For the following processes at 298.15 K: lactulose(aq) + H2O(liq) = D-galactose(aq) + D-fructose(aq), K0 = 128 +/- 10 and delta H0 = 2.21 +/- 0.10 kJ mol-1; alpha-D-melibiose(aq) + H2O(liq) = D-galactose(aq) + D-glucose(aq), K0 = 123 +/- 42 and delta H0 = -0.88 +/- 0.50 kJ mol-1; palatinose(aq) + H2O(liq) = D-glucose(aq) + D-fructose(aq), delta H0 = -4.44 +/- 1.1 kJ mol-1; D-trehalose(aq) + H2O(liq) = 2 D-glucose(aq), K0 = 119 +/- 10 and delta H0 = 4.73 +/- 0.41 kJ mol-1; D-turanose(aq) + H2O(liq) = D-glucose(aq) + D-fructose(aq), delta H0 = -2.68 +/- 0.75 kJ mol-1; and 3-o-beta-D-galactopyranosyl-D-arabinose(aq) + H2O(liq) = D-galactose(aq) + D- arabinose(aq),0H0 = 107 +/- 10 and delta H0 = 2.97 +/- 0.10 kJ mol-1.(ABSTRACT TRUNCATED AT 250 WORDS)

Thermodynamics of hydrolysis of oligosaccharides.

Microcalorimetry has been used to determine enthalpy changes for the hydrolysis of a series of oligosaccharides. High-pressure liquid chromatography was used to determine the extents of reaction and to check for any possible side reactions. The enzyme glucan 1,4-alpha-glucosidase was used to bring about the following hydrolysis reactions: (A) maltose(aq) + H2O(liq) = 2D-glucose(aq); (B) maltotriose(aq) + 2H2O(liq) = 3D-glucose(aq); (C) maltotetraose(aq) + 3H2O(liq) = 4D-glucose(aq); (D) maltopentaose(aq) + 4H2O(liq) = 5D-glucose(aq); (E) maltohexaose(aq) + 5H2O(liq) = 6D-glucose(aq); (F) maltoheptaose(aq) + 6H2O(liq) = 7D-glucose(aq); (G) amylose(aq) + nH2O(liq) = (n + 1) D-glucose(aq); and (H) panose(aq) + 2H2O(liq) = 3D-glucose(aq); (J) isomaltotriose(aq) + 2H2O(liq) = 3D-glucose(aq). The enzyme beta-fructofuranosidase was used for the reactions: (K) raffinose(aq) + H2O(liq) = alpha-D-melibiose(aq) + D-fructose(aq); and (L) stachyose(aq) + H2O(liq) = o-alpha-D-galactopyranosyl-(1----6)- alpha-o-D-galactopyranosyl-(1----6)-alpha-D-glucopyranose + D-fructose(aq). The results of the calorimetric measurements (298.15 K, 0.1 M sodium acetate buffer, pH 4.44-6.00) are: delta H0A = -4.55 +/- 0.10, delta H0B = -9.03 +/- 0.10, delta H0C = -13.79 +/- 0.15, delta H0D = -18.12 +/- 0.10, delta H0E = -22.40 +/- 0.15, delta H0F = -26.81 +/- 0.20, delta H0H = 1.46 +/- 0.40, delta H0J = 11.4 +/- 2.0, delta H0K = -15.25 +/- 0.20, and delta H0L = -14.93 +/- 0.20 kJ mol-1. The enthalpies of hydrolysis of two different samples of amylose were 1062 +/- 20 and 2719 +/- 100 kJ mol-1, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Thermodynamics of industrially-important, enzyme-catalyzed reactions.

The thermodynamics of 10 industrially-important, enzyme-catalyzed reactions are examined. The reactions discussed are: the conversions of penicillin G to 6-amino-penicillinic acid using the enzyme penicillin acylase; starch to glucose using amylases; glucose to fructose using glucose (xylose) isomerase; cellulose to glucose using cellulase; fumaric acid and ammonia to L-aspartic acid using L-aspartase; transcinnamic acid and ammonia to L-phenylalanine using L-phenylalanine ammonia lyase; L-histidine to urocanic acid and ammonia using L-histidine ammonia lyase; lactose to glucose and galactose using lactase; and the reactions catalyzed by amino acylases and proteases. The selection of these processes was based on the economic value of the products and their intrinsic industrial importance. The available thermodynamic properties, such as equilibrium constants, Gibbs energies (delta G degrees), enthalphies (delta H degrees), and heat capacity changes (delta Cp degrees) of these enzyme-catalyzed reactions, are reviewed and summarized. Recommendations are made for future research in this area.

Thermodynamics of the hydrolysis of sucrose.

A thermodynamic investigation of the hydrolysis of sucrose to fructose and glucose has been performed using microcalorimetry and high-pressure liquid chromatography. The calorimetric measurements were carried out over the temperature range 298-316 K and in sodium acetate buffer (0.1 M, pH 5.65). Enthalpy and heat capacity changes were obtained for the hydrolysis of aqueous sucrose (process A): sucrose(aq) + H2O(liq) = glucose(aq) + fructose (aq). The determination of the equilibrium constant required the use of a thermochemical cycle calculation involving the following processes: (B) glucose 1-phosphate2-(aq) = glucose 6-phosphate2-(aq); (C) sucrose(aq) + HPO4(2-)(aq) = glucose 1-phosphate2-(aq) + fructose(aq); and (D) glucose 6-phosphate2-(aq) + H2O(liq) = glucose(aq) + HPO4(2-)(aq). The equilibrium constants determined at 298.15 K for processes B and C are 17.1 +/- 1.0 and 32.4 +/- 3.0, respectively. Equilibrium data for process D was obtained from the literature, and in conjunction with the data for processes B and C, used to calculate a value of the equilibrium constant for the hydrolysis of aqueous sucrose. Thus, for process A, delta G0 = -26.53 +/- 0.30 kJ mol-1, K0 = (4.44 +/- 0.54) x 10(4), delta H0 = -14.93 +/- 0.16 kJ mol-1, delta So = 38.9 +/- 1.2 J mol-1 K-1, and delta CoP = 57 +/- 14 J mol-1 K-1 at 298.15 K. Additional thermochemical cycles that bear upon the accuracy of these results are examined.

A calorimetric and equilibrium investigation of the hydrolysis of lactose.

The thermodynamics of the hydrolysis of lactose to glucose and galactose have been investigated using both high pressure liquid chromatography and heat-conduction microcalorimetry. The reaction was carried out over the temperature range 282-316 K and in 0.1 M sodium acetate buffer at a pH of 5.65 using the enzyme beta-galactosidase to catalyze the reaction. For the process lactose(aq) + H2O(liq) = glucose(aq) + galactose(aq), delta G0 = -8.72 +/- 0.20 kJ.mol-1, K0 = 34 +/- 3, delta H0 = 0.44 +/- 0.11 kJ.mol-1, delta S0 = 30.7 +/- 0.8 J.mol-1.K-1, and delta Cop = 9 +/- 20 J.mol-1.K-1 at 298.15 K. The standard state is the hypothetical ideal solution of unit molality. Thermochemical cycle calculations using enthalpies of combustion and solution, entropies, solubilities, activity coefficients, and apparent molar heat capacities have also been performed. These calculations indicate large discrepancies which are attributable primarily to errors in literature data on the enthalpies of combustion and/or third law entropies of the crystalline forms of the substrates.

Thermodynamics of hydrolysis of disaccharides. Cellobiose, gentiobiose, isomaltose, and maltose.

The thermodynamics of the enzymatic hydrolysis of cellobiose, gentiobiose, isomaltose, and maltose have been studied using both high pressure liquid chromatography and microcalorimetry. The hydrolysis reactions were carried out in aqueous sodium acetate buffer at a pH of 5.65 and over the temperature range of 286 to 316 K using the enzymes beta-glucosidase, isomaltase, and maltase. The thermodynamic parameters obtained for the hydrolysis reactions, disaccharide(aq) + H2O(liq) = 2 glucose(aq), at 298.15 K are: K greater than or equal to 155, delta G0 less than or equal to -12.5 kJ mol-1, and delta H0 = -2.43 +/- 0.31 kJ mol-1 for cellobiose; K = 17.9 +/- 0.7, delta G0 = -7.15 +/- 0.10 kJ mol-1 and delta H0 = 2.26 +/- 0.48 kJ mol-1 for gentiobiose; K = 17.25 +/- 0.7, delta G0 = -7.06 +/- 0.10 kJ mol-1, and delta H0 = 5.86 +/- 0.54 kJ mol-1 for isomaltose; and K greater than or equal to 513, delta G0 less than or equal to -15.5 kJ mol-1, and delta H0 = -4.02 +/- 0.15 kJ mol-1 for maltose. The standard state is the hypothetical ideal solution of unit molality. Due to enzymatic inhibition by glucose, it was not possible to obtain reliable values for the equilibrium constants for the hydrolysis of either cellobiose or maltose. The entropy changes for the hydrolysis reactions are in the range 32 to 43 J mol-1 K-1; the heat capacity changes are approximately equal to zero J mol-1 K-1. Additional pathways for calculating thermodynamic parameters for these hydrolysis reactions are discussed.

Thermodynamics of the conversion of penicillin G to phenylacetic acid and 6-aminopenicillanic acid.

The thermodynamics of the enzymatic conversion (penicillin acylase) of aqueous penicillin G to phenylacetic acid and 6-aminopenicillanic acid have been studied using both high-pressure liquid-chromatography and microcalorimetry. The reaction was carried out in aqueous phosphate buffer over the pH range 6.0-7.6, at ionic strengths from 0.10 to 0.40 mol kg-1, and at temperatures from 292 to 322 K. The data have been analyzed using a chemical equilibrium model with an extended Debye-Hückel expression for the activity coefficients. For the reference reaction, penicillin G- (aq) + H2O(l) = phenylacetic acid-(aq) + 6-aminopenicillanic acid-(aq) + H+ (aq), the following parameters have been obtained: K = (7.35 +/- 1.5) X 10(-8) mol kg-1, delta G0 = 40.7 +/- 0.5 kJ mol-1, delta H0 = 29.7 +/- 0.6 kJ mol-1, and delta C0p = -240 +/- 50 J mol-1 K-1 at 298.15 K and at the thermochemical standard state. The extent of reaction for the overall conversion is highly dependent upon the pH.

Thermodynamics of isomerization reactions involving sugar phosphates.

Thermodynamics of isomerization reactions involving sugar phosphates have been studied using heat-conduction microcalorimetry. For the process glucose 6-phosphate2-(aqueous) = fructose 6-phosphate2- (aqueous), K = 0.285 +/- 0.004, delta Go = 3.11 +/- 0.04 kJ.mol-1, delta Ho = 11.7 +/- 0.2 kJ.mol-1, and delta Cop = 44 +/- 11 J.mol-1.K-1 at 298.15 K. For the process mannose 6-phosphate2- (aqueous) = fructose 6-phosphate2- (aqueous), K = 0.99 +/- 0.05, delta Go = 0.025 +/- 0.13 kJ.mol-1, delta Ho = 8.46 +/- 0.2 kJ.mol-1, and delta Cop = 38 +/- 25 J.mol-1.K-1 at 298.15 K. The standard state is the hypothetical ideal solution of unit molality. An approximate result (-14 +/- 5 kJ.mol-1) was obtained for the enthalpy of isomerization of ribulose 5-phosphate (aqueous) to ribose 5-phosphate (aqueous). The data from the literature on isomerization reactions involving sugar phosphates have been summarized, adjusted to a common reference state, and examined for trends and relationships to each other and to other thermodynamic measurements. Estimates are made for thermochemical parameters to predict the state of equilibrium of the several isomerizations considered herein.

Thermodynamics of hydrolysis of sugar phosphates.

Thermodynamics of the enzyme-catalyzed (alkaline phosphatase, EC 3.1.3.1) hydrolysis of glucose 6-phosphate, mannose 6-phosphate, fructose 6-phosphate, ribose 5-phosphate, and ribulose 5-phosphate have been investigated using microcalorimetry and, for the hydrolysis of fructose 6-phosphate, chemical equilibrium measurements. Results of these measurements for the processes sugar phosphate2- (aqueous) + H2O (liquid) = sugar (aqueous) + HPO2++-(4) (aqueous) at 25 degrees C follow: delta Ho = 0.91 +/- 0.35 kJ.mol-1 and delta Cop = -48 +/- 18 J.mol-1.K-1 for glucose 6-phosphate; delta Ho = 1.40 +/- 0.31 kJ.mol-1 and delta Cop = -46 +/- 11 J.mol-1.dK-1 for mannose 6-phosphate; delta Go = -13.70 +/- 0.28 kJ.mol-1, delta Ho = -7.61 +/- 0.68 kJ.mol-1, and delta Cop = -28 +/- 42 J.mol-1.K-1 for fructose 6-phosphate; delta Ho = -5.69 +/- 0.52 kJ.mol-1 and delta Cop = -63 +/- 37 J.mol-1.K-1 for ribose 5-phosphate; and delta Ho = -12.43 +/- 0.45 kJ.mol-1 and delta Cop = -84 +/- 30 J.mol-1.K-1 for the hydrolysis of ribulose 5-phosphate. The standard state is the hypothetical ideal solution of unit molality. Estimates are made for the equilibrium constants for the hydrolysis of ribose and ribulose 5-phosphates. The effects of pH, magnesium ion concentration, and ionic strength on the thermodynamics of these reactions are considered.

Thermodynamics of carbohydrate isomerization reactions. The conversion of aqueous allose to psicose.

The thermodynamics of the conversion of aqueous D-psicose to D-allose has been investigated using high-pressure liquid chromatography. The reaction was carried out in phosphate buffer at pH 7.4 over the temperature range 317.25-349.25 K. The following results are obtained for the conversion process at 298.15 K: DeltaG degrees = - 1.41 +/- 0.09 kJ mol(-1), DeltaH degrees = 7.42 +/- 1.7 kJ mol(-1), and DeltaC(p) degrees = 67 +/- 50 J mol(-1) K(-1). An approximate equilibrium constant of 0.30 is obtained at 333.15 K for the conversion of aqueous D-psicose to D-altrose. Available thermodynamic data for isomerization reactions involving aldohexoses and aldopentoses are summarized.

Thermodynamics of the conversion of aqueous L-aspartic acid to fumaric acid and ammonia.

The thermodynamics of the conversion of aqueous L-aspartic acid to fumaric acid and ammonia have been investigated using both heat conduction microcalorimetry and high-pressure liquid chromatography. The reaction was carried out in aqueous phosphate buffer over the pH range 7.25-7.43, the temperature range 13-43 degrees C, and at ionic strengths varying from 0.066 to 0.366 mol kg(-1). The following values have been found for the conversion of aqueous L-aspartateH- to fumarate2- and NH4+ at 25 degrees C and at zero ionic strength: K = (1.48 +/- 0.10) x 10(-3), DeltaG degrees = 16.15 +/- 0.16 kJ mol(-1), DeltaH degrees = 24.5 +/- 1.0 kJ mol(-1), and DeltaC(p) degrees = -147 +/- 100 J mol(-1) K(-1). Calculations have also been performed which give values of the apparent equilibrium constant for the conversion of L-aspartic acid to fumaric acid and ammonia as a function of temperature, pH and ionic strength.

High Precision Microcalorimetry: Apparatus, Procedures, and Biochemical Applications.

Apparatus and procedures used for high-precision microcalorimetric measurements are described. The calorimeter is of the heat-conduction type and utilizes semi-conductor thermoelectric modules. The bicompartmental reaction vessel is made of high-density polyethylene and holds about 0.5 mL of solution in each compartment. Imprecision of heat measurement is 0.2 percent when measuring 300 mJ of heat produced by a rapid chemical reaction. Three microcalorimeters are operated simultaneously using a microcomputer and a data acquisition system. Thermochemical and kinetic applications are described. The acquisition of data from an isoperibol solution calorimeter is also described.

An investigation of the equilibria between aqueous ribose, ribulose, and arabinose.

The thermodynamics of the equilibria between aqueous ribose, ribulose, and arabinose were investigated using high-pressure liquid chromatography and microcalorimetry. The reactions were carried out in aqueous phosphate buffer over the pH range 6.8-7.4 and over the temperature range 313.15-343.75 K using solubilized glucose isomerase with either Mg(NO3)2 or MgSO4 as cofactors. The equilibrium constants (K) and the standard state Gibbs energy (delta G degrees) and enthalpy (delta H degrees) changes at 298.15 K for the three equilibria investigated were found to be: ribose(aq) = ribulose(aq) K = 0.317, delta G degrees = 2.85 +/- 0.14 kJ mol-1, delta H degrees = 11.0 +/- 1.5 kJ mol-1; ribose(aq) = arabinose(aq) K = 4.00, delta G degrees = -3.44 +/- 0.30 kJ mol-1, delta H degrees = -9.8 +/- 3.0 kJ mol-1; ribulose(aq) = arabinose(aq) K = 12.6, delta G degrees = -6.29 +/- 0.34 kJ mol-1, delta H degrees = -20.75 +/- 3.4 kJ mol-1. Information on rates of the above reactions was also obtained. The temperature dependencies of the equilibrium constants are conveniently expressed as R in K = -delta G degrees 298.15/298.15 + delta H degrees 298.15[(1/298.15)-(1/T)] where R is the gas constant (8.31441 J mol-1 K-1) and T the thermodynamic temperature.