Biomedical subjects

T Flatmark

Publications and source records attributed to T Flatmark.

At least 109 records · Page 6Linked to original sources

PubMed1985

Isolation from the microsomal fraction of rat liver of a subfraction highly enriched in uncoated endocytic vesicles with high H+-ATPase activity and a 50 kDa phosphoprotein.

A subcellular fraction, highly enriched in uncoated vesicles (UCV) with high H+-ATPase (EC 3.6.1.34) activity, was isolated from the crude microsomal fraction of rat liver homogenates by discontinuous sucrose gradient centrifugation. The UCV fraction, recovered at the interface of sucrose density 1.08 and 1.10 g/ml, was shown morphologically to be a mixture of small, smooth-surfaced univesicular and a few multivesicular structures. A permeable anion (e.g. chloride) was required for internal acidification, indicating an electroneutral proton pump. Specific inhibitors of anion transport (pyridoxal 5'-phosphate and 4-acetamide-4'-isothiocyanostilbene-2,2'-disulfonic acid) totally inhibit proton translocation. The proton pump activity was insensitive to oligomycin, but was completely inhibited by about 5 microM of the tridentate bathophenanthroline chelate of Fe(II). The activity was also inhibited 100% by low concentrations of the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone, the proton conduction inhibitor N,N'-dicyclohexylcarbodiimide and the ionophore monensin. The UCV fraction contained 2 proteins of Mr 50000 (major) and 54000 (minor) which were phosphorylated by an endogenous cyclic nucleotide- and Ca2+-independent protein kinase.

Adenosine Triphosphatases↗

PubMed1985

The assignment of the Ca2+-ATPase activity of chromaffin granules to the proton translocating ATPase.

CaATP is shown to function as a substrate for the proton translocating ATPase of chromaffin granule ghosts at concentrations which are comparable to that of MgATP. Using the initial rate of the proton pump activity as the measure (delta pH/delta t), an apparent Km-value of 139 +/- 8 microM was estimated for CaATP and 59 +/- 3 microM for MgATP. The maximal rate was markedly higher with MgATP than with CaATP, partly due to an inhibition of the hydrolytic activity at the higher concentrations of CaATP. The proton pump activity with CaATP was inhibited by N-ethylmaleimide and N,N'-dicyclohexylcarbodiimide at concentrations similar to that found for MgATP. No inhibition was observed with sodium vanadate in the concentration range 0-15 microM. Calmodulin and trifluoperazine had no effect on the overall ATPase activity with CaATP. These findings establish this activity as an intrinsic property of the chromaffin granules, i.e., linked to the H+-ATPase. No evidence was obtained for the presence of a Ca2+-translocating ATPase [Ca2+ + Mg2+)-ATPase) in the chromaffin granules.

Adenosine Triphosphatases↗

PubMed1984

The content of long-chain free fatty acids and their effect on energy transduction in chromaffin granule ghosts.

Highly purified chromaffin granule ghosts contain about 100 (89.1-110.2, n = 8) nmol of long-chain free fatty acids/mg of protein. About 40% of the fatty acids were mono- and polyunsaturated, of which arachidonate accounted for approximately 20%. Incubation with fatty acid-free bovine serum albumin selectively depleted the ghosts of unsaturated fatty acids. The effect of a series of long-chain fatty acids on energy coupling was studied in resealed chromaffin granule ghosts energized by MgATP. Their effectiveness in inhibiting the generation of the transmembrane proton concentration gradient and transmembrane potential was found to be correlated with their chain length and degree of unsaturation, all-cis-5,8,11,14-icosatetraenoate (arachidonate) and all-cis-5,8,11,14,-17-icosapentaenoate being most effective. Arachidonate was also found to dissipate an artificially generated pH gradient. Treatment of chromaffin granule ghosts with bovine serum albumin increased the transmembrane proton concentration gradient that could be generated by MgATP. This enhanced response to MgATP is presumably due to the removal of free fatty acids from the ghosts. Considering the potency of arachidonate and other unsaturated fatty acids, as well as their high endogenous content, these "uncoupling agents" should be accounted for when isolated chromaffin granules are used for in vitro studies on energy coupling.

Adenosine Triphosphatases↗

PubMed1984

Some aspects of the phosphorylation of phenylalanine 4-monooxygenase by a calcium-dependent and calmodulin-dependent protein kinase.

A calmodulin-dependent protein kinase purified from liver catalyzed the incorporation of up to 0.7 mol of phosphate per mol subunit of phenylalanine 4-monooxygenase. The phosphorylation was accompanied by a proportional increase in the hydroxylase activity. The reaction was Ca2+-dependent and was inhibited by physiological concentrations of phenylalanine. Phenylalanine 4-monooxygenase was also a substrate for the cGMP-dependent protein kinase, but in this system phenylalanine stimulated the rate of phosphorylation to a similar extent as that observed in the reaction catalyzed by cAMP-dependent protein kinase. The hydroxylase was not a substrate for phosphorylase kinase. The calmodulin-dependent reversal of the kinase reaction in the presence of MgADP, was also inhibited by phenylalanine. Since the kinetics of the reverse reaction was the same using 32P-hydroxylase phosphorylated by calmodulin-dependent and cAMP-dependent kinases, it is likely that both kinases phosphorylate the same site on the enzyme. This conclusion was further supported by peptide mapping of tryptic and peptic digests of 32P-hydroxylase, which revealed one major phosphopeptide with enzyme phosphorylated by either kinase. The Ca2+-dependent and calmodulin-dependent phosphorylation described above may mediate the increased phosphorylation of the hydroxylase [Garrison, J. C., Johnsen, D. E., and Campanile, C. P. (1984) J. Biol. Chem. 259, 3283-3292] and its increased activity [Fisher, M. J., Santana, M. A., and Pogson, C. I. (1984) Biochem. J. 219, 87-90] recently observed in hepatocytes exposed to Ca2+-elevating agents.

Animals↗

PubMed1984

The effect of ligands of phenylalanine 4-monooxygenase on the cAMP-dependent phosphorylation of the enzyme.

The rate of phosphorylation of phenylalanine 4-monooxygenase by the cAMP-dependent protein kinase was found to be under substrate-directed regulation. Thus, L-phenylalanine made the hydroxylase a better substrate for the kinase, whereas the cofactor l-erythro-5,6,7,8-tetrahydrobiopterin (BH4) was a negative effector. The dephosphorylation of the enzyme by the kinase in the presence of high concentrations of MgADP was also stimulated by phenylalanine and inhibited by BH4. A kinetic analysis indicated that the effects of phenylalanine and BH4 were mediated by distinct sites coupled by a free energy of 3.2 kJ X mol-1. Among the ligands tested, only phenylalanine and BH4 affected the phosphorylation of the hydroxylase at physiologically relevant concentrations. Whereas higher concentrations of several naturally occurring or synthetic amino acids acted like phenylalanine, the widely used synthetic cofactor 6,7-dimethyltetrahydropterin did not mimic the effect of BH4. Less phenylalanine was required to activate the phosphorylated hydroxylase (0.9 mol of phosphate/subunit) than the dephosphorylated enzyme (0.07 mol of phosphate/subunit). This was true whether BH4 was present or not. In conclusion, the substrate phenylalanine makes the hydroxylase more prone to cAMP-dependent phosphorylation, which in turn sensitizes the enzyme towards allosteric activation by phenylalanine. The joint operation of these mechanisms in vivo would increase the efficiency with which phenylalanine controls the activity of the enzyme.

Adenosine Diphosphate↗

PubMed1984

Demonstration and partial purification of lactoperoxidase from human colostrum.

A peroxidase with stability, chromatographic and immunoreactive properties similar to that of bovine lactoperoxidase has been partly purified from human colostrum. Hydrophobic interaction chromatography on Phenyl-Sepharose C1-4B gave a 10-fold purification with an apparent recovery of about 45%. The enzyme was quantitatively and specifically adsorbed to beads of anti-lactoperoxidase (bovine)-Protein A-Sepharose. No adsorption of the enzyme was observed on immunoadsorbent columns prepared with high-titre polyclonal antibodies raised against human myeloperoxidase and human eosinophile peroxidase.

Chromatography, Affinity↗

PubMed1984

Increase in mitochondrial content of long-chain acyl-CoA in brown adipose tissue during cold-acclimation.

The mitochondrial content of long-chain acyl-CoA esters in the brown adipose tissue of guinea pigs increased 3.5-fold from a level of 92 +/- 17 pmol per mg protein (+/- S.E.; n = 7) in the control animals adapted at 22 degrees C to a new steady-state level of 328 +/- 20 pmol per mg protein (+/- S.E.; n = 46) after 10 days of cold-acclimation (5 degrees C). These low values of long-chain acyl-CoA species and the slow adaptive response for their increase do not support the proposal (Cannon, B., Sindin, U. and Romert, L. (1977) FEBS Lett. 4, 43-46) that the fatty acid CoA-esters have a physiological function in the regulation of the H+ (or OH-) permeability of the mitochondrial inner membrane. Experimental evidence is presented supporting the proposal that the long-chain acyl-CoA species are largely confined to the cytosolic side of the inner membrane. The activity of the adenine nucleotide translocase, as estimated at 25 degrees C in the reverse direction, was found to increase 5-fold upon depletion of the mitochondria of fatty acids (free and esterified) by preincubation with bovine serum albumin. The presence of potent inhibitors, i.e., long-chain acyl-CoA species, of adenine nucleotide translocation in brown adipose tissue of thermogenically active animals further supports the conclusion that ATP hydrolyzing mechanisms contribute insignificantly to long-term thermogenesis. The low values of long-chain acyl-CoA hydrolase (EC 3.1.2.1) activity, as measured in intact mitochondria and on a mitochondrial matrix fraction (i.e., 1.6 nmol X min-1 per mg protein), do not support the proposal that the hydrolase activity plays a significant role in the loose-coupling of brown adipose tissue mitochondria, either by a futile cycle mechanism or promoted by free fatty acid-induced uncoupling.

Acyl Coenzyme A↗

PubMed1984

Enhancement of long-chain acyl-CoA hydrolase activity in peroxisomes and mitochondria of rat liver by peroxisomal proliferators.

The present study has confirmed previous findings of long-chain acyl-CoA hydrolase activities in the mitochondrial and microsomal fractions of the normal rat liver. In addition, experimental evidence is presented in support of a peroxisomal localization of long-chain acyl-CoA hydrolase activity. (a) Analytical differential centrifugation of homogenates from normal rat liver revealed that this activity (using palmitoyl-CoA as the substrate) was also present in a population of particles with an average sedimentation coefficient of 6740 S, characteristic of peroxisomal marker enzymes. (b) The subcellular distribution of the hydrolase activity was greatly affected by administration of the peroxisomal proliferators clofibrate and tiadenol. The specific activity was enhanced in the mitochondrial fraction and in a population of particles with an average sedimentation coefficient of 4400 S, characteristic of peroxisomal marker enzymes. Three populations of particles containing lysosomal marker enzymes were found by analytical differential centrifugation, both in normal and clofibrate-treated rats. Our data do not support the proposal that palmitoyl-CoA hydrolase and acid phosphatase belong to the same subcellular particles. In livers from rats treated with peroxisomal proliferators, the specific activity of palmitoyl-CoA hydrolase was also enhanced in the particle-free supernatant. Evidence is presented that this activity at least in part, is related to the peroxisomal proliferation.

Animals↗

PubMed1984

The copper-enzyme dopamine beta-monooxygenase: studies on the ability of several metals to inhibit the enzyme activity and to replace the copper.

The ability of several metals to inhibit dopamine beta-monooxygenase was measured and compared with their ability to compete with the binding of 64Cu to the water-soluble form of the bovine adrenal enzyme at pH 6.0. In the presence of an optimal concentration of copper (0.5 microM in the present assay system), an inhibition was observed upon addition of Hg(II), Zn(II), or Ni(II). Only a small fraction of the inhibition with these metals may be due to uncoupling of electron transport from hydroxylation. Preincubation of these metals with the Cu-depleted apoenzyme before addition of copper, revealed a stronger inhibition than if copper was added before the other metals. Hg(II), Zn(II), and Ni(II) also compete with the binding of 64Cu(II) to the protein. Hg(II) was the most effective and Ni(II) the least effective of these metals, both with respect to inhibition of the enzyme activity and to prevent the binding of 64Cu(II). Competition experiments on the binding of Zn(II) and 64Cu in the presence and absence of ascorbate, indicated i) a similar affinity of Cu(I) and Cu(II) to the native enzyme, and ii) a more rapid binding of Cu(I) than Cu(II) to the Cu-depleted and Zn-containing enzyme. Al(III), Fe(II), Mg(II), Mn(II), Co(II), Cd(II), and Pb(II) neither inhibited the enzyme activity nor competed with the binding of 64Cu(II) to the protein (Fe(II) was not tested for binding). Of those metals cited above only Cu(II)/Cu(I) was able to reactivate the apoenzyme.

Adrenal Glands↗

PubMed1983

Evidence for a negative modulating effect of erucic acid on the peroxisomal beta-oxidation enzyme system and biogenesis in rat liver.

In male rats fed a 30 cal% high-erucic acid (22:1 (13) (cis] rapeseed oil diet for 4 weeks a transient, small increase in peroxisomal beta-oxidation was observed in liver homogenates and isolated peroxisomes after approximately 1 week. Morphometric analysis revealed a progressive decrease in the average size of the liver peroxisomes (by approx. 20%), as well as their volume fraction (by more than 40%). A negative dose-response was observed when peroxisomal beta-oxidation was determined in animals fed rapeseed oil diets with a variable content of erucic acid. When erucic acid was given as subcutaneous injections the peroxisomal beta-oxidation decreased, in both liver homogenates and isolated peroxisomes. In contrast to recent proposals, our findings indicate that the amount of erucic acid cannot account for the small increase in peroxisomal beta-oxidation observed in the liver of rats adapted to a diet containing 30 cal% rapeseed oil with a high content of this fatty acid. In fact, by the selected criteria erucic acid seems to have a negative modulating effect on both the peroxisomal beta-oxidation enzyme system and biogenesis.

3-Hydroxyacyl CoA Dehydrogenases↗

PubMed1983

Acyl-CoA synthetase activity of rat heart mitochondria. Substrate specificity with special reference to very-long-chain and isomeric fatty acids.

The acyl-CoA synthetase (acid: CoA ligase (AMP-forming), EC 6.2.1.3) activity of rat heart has been measured in fatty acid-depleted fractions of mitochondria, microperoxisomes and microsomes. The assay was based on (i) the measurement of the reaction product AMP by high-performance liquid chromatography or (ii) a coupled reaction in which the intramitochondrial (matrix) CoASH is the final acyl acceptor and the redox state of the flavoproteins in the acyl-CoA dehydrogenase pathway is used to determine the intramitochondrial level of acyl-CoA. This spectrophotometric method was also used to estimate the 'outer' carnitine long-chain acyltransferase (palmitoyl-CoA:L-carnitine O-palmitoyltransferase, EC 2.3.1.21) activity. Comparison of the distribution of long-chain acyl-CoA synthetase activity and marker enzymes in the various subcellular fractions revealed that the synthetase activity is exclusively localized in the mitochondrial fraction. Experimental evidence is presented in support of the conclusion that the chain-length specificity of saturated and monounsaturated fatty acids (16:1-22:1) for the acyl-CoA synthetase is mainly determined by the availability of the fatty acid at the active site, which is largely determined by the affinity of binding of fatty acids to the bulk phase of the mitochondrial phospholipids. Among the 22:1 isomers, 22:1(11) (cis) (cetoleic acid) revealed a slightly higher activity (1.4-fold) than 22:1(13) (cis) (erucic acid). The polyunsaturated fatty acids tested were rather poor substrates. Using isolated intact mitochondria and 16:0 or 22:1(13) (cis) as the substrates, it was found that the initial rate of the 'outer' long-chain acyltransferase activity was approximately four times higher than that of the long-chain acyl-CoA synthetase. The data support the hypothesis that the long-chain acyl-CoA synthetase reaction is rate-limiting in the sequence of coupled reactions leading to beta-oxidation in the mitochondrial matrix.

Animals↗

PubMed1983

Dopamine beta-monooxygenase. Binding to apoenzyme and rapid exchange in holoenzyme of 64Cu studied with high-performance size-exclusion gel chromatography.

The binding of 64Cu to the water-soluble form of dopamine beta-monooxygenase from bovine adrenal medulla was studied in reconstitution and exchange experiments using high-performance size-exclusion gel chromatography. The reconstitution experiments provide evidence for a specific binding of four copper atoms/enzyme tetramer using either Cu(I) or Cu(II), but some weaker copper-binding sites were observed in the presence of a large excess of copper. The exchanges of both Cu(I) and Cu(II) in this protein are so rapid that exact half-lives for the exchange reactions can not be obtained by the present method. The results indicate, however, that the half-life for the exchange of the enzyme-bound copper in the holoenzyme with a twofold excess of 64Cu(II) at pH 6.1 was about 1 min, whereas the exchange of Cu(I) measured at similar conditions with ascorbate present, was complete in 1 min. This is by far the most rapid exchange reported for any copper-protein, and the results points to a unique copper-binding site in this enzyme.

Adrenal Medulla↗

PubMed1983

Time course of changes in nociception after 5,6-dihydroxytryptamine lesions of descending 5-HT pathways.

Intrathecal injection of 5,6-dihydroxytryptamine (5,6-DHT) in rats produced selective lesions of the descending 5-HT pathways. Spinal 5-HT levels gradually fell to less than 10% of controls within 10 days of 5,6-DHT administration with no recovery evident within 4 weeks. The uptake of 14C-5-HT into crude spinal synaptosomes was similarly reduced. The uptake of 3H-NA into spinal synaptosomes was unaffected, as was the uptake of 14C-5-HT and 3H-NA into cortical synaptosomes. Following 5,6-DHT, tail-flick latencies were reduced by 20-30% during the first post-injection week, but returned to control levels during the second week. Intrathecal or systemic administration of the 5-HT receptor antagonist metergoline significantly reduced latencies of normal rats and of 5,6-DHT treated rats tested after the second week when the response was normalized. Metergoline did not, however, further reduce the latencies of lesioned rats during the first post-injection week. It is concluded that functional adaptation involving 5-HT neurotransmission compensated for the selective lesion of descending 5-HT pathways induced by 5,6-DHT.

5,6-Dihydroxytryptamine↗

PubMed1982

Hydrodynamic parameters and isolation of mitochondria, microperoxisomes and microsomes of rat heart.

1. Analytical differential centrifugation of rat heart homogenates revealed a single population of mitochondria and microperoxisomes. Using cytochrome c oxidase, malate dehydrogenase and amine oxidase as mitochondrial marker enzymes, the s-value of mitochondria was estimated to s = 10326 +/- 406 S (average for the three marker enzymes). The s-value of microperoxisomes was found to be s = 1381 +/- 40 S using catalase as the marker enzyme. The s-value for the two organelles did not change significantly when the isoosmotic sucrose medium was substituted by an isoosmotic mannitol medium. 2. Analytical differential centrifugation revealed a polydispercity of the microsomal fraction using glucose-6-phosphatase and NADPH-cytochrome c reductase as the marker enzymes. The s-values were found to be sH1 = 1569 +/- 412 S (NADPH-cytochrome c reductase), sH2 = 1195 +/- 400 S (glucose-6-phosphatase) and sL = 153 +/- 28 S (NADPH-cytochrome c reductase and glucose-6-phosphatase). The recovery of marker enzymes in the isolated subcellular fractions was in the range of 84-94%. 3. When the mitochondrial and microperoxisomal fractions were subjected to isopycnic gradient centrifugation, using a self-generating gradient of polyvinylpyrrolidone-coated colloidal silica particles (Percoll) in 0.25 M sucrose medium, buoyant densities of 1.10 g/cm3 (main fraction of mitochondria) and 1.06 g/cm3 (main fraction of microperoxisomes) were obtained. The density gradient centrifugation separated microperoxisomes from contaminating lysosomes of high specific activity in acid phosphatase. A value 1.04 g/cm3 was found for the density of the microsomal fraction. 4. Based on the estimated s-values, an optimal procedure is described for the isolation of mitochondrial and microperoxisomal fractions from rat heart muscle.

Animals↗

PubMed1982

Inhibition of N-ethylmaleimide of the MgATP-driven proton pump of the chromaffin granules.

The thiol reagent N-ethylmaleimide (NEM) completely inhibits the proton pump activity of the H+-ATPase in chromaffin granule 'ghosts' at concentrations which only partly (approximately 20%) inhibit the Mg2+-dependent ATP hydrolysis. Half-maximal inhibition was obtained at approximately 13 microM NEM as compared to 18 microM for the classical proton channel inhibitor N,N'-dicyclohexylcarbodiimide (DCCD), and the apparent stoichiometry of the inhibitors at complete inhibition was NEM : DCCD congruent to 1 : 2. HIgh concentrations of NEM (greater than 100 microM) induce a dissipation of the transmembrane potential generated by MgATP. These findings establish NEM as a valuable proton channel inhibitor in chromaffin granules and explain the rather complex effect of NEM previously reported for catecholamine accumulation in this organelle.

Adenosine Triphosphatases↗

PubMed1982

Microperoxisomes and mitochondria of brown adipose tissue. Hydrodynamic parameters, isolation and capacity of long-chain fatty acid oxidation.

1. Analytical differential centrifugation of brown adipose tissue homogenates from cold-acclimated guinea pigs revealed a polydispersity of both mitochondria and peroxisomes, with at least two populations of each organelle. The estimated values were sH = 16685+/-4220 S and sL = 4792+/-951 S (mitochondria), and sH = 3364+/-1706 S and sL = 889+/-177 S (peroxisomes). Based on these s values, an optimal procedure is described for the isolation of subcellular fractions enriched in mitochondria and peroxisomes, respectively. 2. When the mitochondrial and peroxisomal fractions were subjected to isopycnic gradient centrifugation on a self-generating gradient of poly(vinylpyrrolidone)-coated colloidal silica particles (Percoll) in 0.25 M sucrose medium, buoyant densities of about 1.11 g/cm3 (main fraction of mitochondria) and 1.07 g/cm3 (main fraction of peroxisomes) were obtained. A value of 1.06 g/cm3 was found for the microsomal fraction. 3. The main peroxisomal fraction, isolated by gradient centrifugation, did not reveal any significant oxidation of palmitoyl-CoA as measured by conventional polarographic technique, whereas a small rate of oxidation (about 2.7+/-0.2 nmol/min per mg peroxisomal protein) was observed when measured as NAD+ reduction. This rate contributes no more than 1% of the mitochondrial oxidation of this fatty acid and it is, therefore, concluded that peroxisomal oxidation of the predominant long-chain fatty acids found in this tissue does not make a quantitatively significant contribution to fatty acid oxidation.

Acyl Coenzyme A↗

PubMed1982

Spectral properties of the oestrogen-induced rat uterus peroxidase II and some of its derivatives.

1. The visible absorption spectrum of peroxidase II, isolated from the uterine tissue of oestradiol-treated rats, and some of its derivatives were recorded. The spectral properties of this enzyme are very similar to eosinophile peroxidase and lactoperoxidase, suggesting that these enzymes may have a similar form of haem as prosthetic group. 2. The uterine peroxidase is modified upon interaction with H2O2 and the difference spectrum of this modified enzyme is similar to that of complex II of lactoperoxidase. The modified enzyme was found to revert spontaneously to the native enzyme at rates which depended on the concentration of free enzyme and H2O2.

Animals↗

PubMed1982

Phenylalanine 4-monooxygenase from bovine and rat liver: some physical and chemical properties.

Phenylalanine 4-monooxygenase was purified from bovine liver using a modification of the procedure developed for the rat liver enzyme (Shiman, R., Gray, D.W., and Pater, A. 1979. J. Biol. Chem. 254: 11300-11306). The enzyme preparation appeared essentially homogeneous on polyacrylamide gel electrophoresis under non-denaturing conditions. Electrophoresis in the presence of dodecyl sulfate revealed that about 95% of the protein had a mobility corresponding to Mr = 51,000. The remaining 5% was recovered in two minor bands corresponding to Mr of about 35,000 and 15,000 and is likely to result from limited proteolysis of the native enzyme with dissociation of the fragments on denaturation by detergent. The enzyme comigrated with the rat liver enzyme on polyacrylamide gel electrophoresis in both systems studied. No significant difference was observed between the amino acid composition of the bovine and rat liver enzyme, in the reactivity of their sulfhydryl groups or in their iron content (i.e. 1.5-3.0 iron atoms per peptide chain of Mr = 50,000). Both enzymes contained less than 0.01 copper atom per peptide chain. The enzymes were inhibited in a similar manner by the chelator bathophenanthroline disulfonate (selective for iron and copper), but not by bathocuproine disulfonate (specific for copper). The results indicate that the bovine and rat liver enzymes are closely similar and that iron, but not copper, is essential for enzyme activity. High performance size-exclusion liquid chromatography revealed that both native enzymes exist in different oligomeric forms, but further studies are required to understand the physicochemical basis for this phenomenon.

Amino Acid Sequence↗