Biomedical subjects

J Preiss

Publications and source records attributed to J Preiss.

At least 19 recordsLinked to original sources

PubMed2006

ADPglucose pyrophosphorylase's N-terminus: structural role in allosteric regulation.

We studied the functional role of the Escherichia coli ADPglucose pyrophosphorylase's N-terminus in allosteric regulation, and the particular effects caused by its length. Small truncated mutants were designed, and those lacking up to 15-residues were active and highly purified for further kinetic analyses. Ndelta3 and Ndelta7 did not change the kinetic parameters with respect to the wild-type. Ndelta11 and Ndelta15 enzymes were insensitive to allosteric regulation and highly active in the absence of the activator. Co-expression of two polypeptides corresponding to the N- and C-termini generated an enzyme with activation properties lower than those of the wild-type [C.M. Bejar, M.A. Ballicora, D.F. Gómez Casati, A.A. Iglesias, J. Preiss, The ADPglucose pyrophosphorylase from Escherichia coli comprises two tightly bound distinct domains, FEBS Lett. 573 (2004) 99-104]. Here, we characterized a Ndelta15 co-expression mutant, in which the allosteric regulation was restored to wild-type levels. Unusual allosteric effects caused by either an N-terminal truncation or co-expression of individual domains may respond to structural changes favoring an up-regulated or a down-regulated conformation rather than specific activator or inhibitor sites' disruption.

Allosteric Regulation↗

PubMed2006

Moderate hyperhomocysteinemia in patients treated for epilepsy.

Homocystein (Hcy) is regarded as a neuroexcitatory substance, which is therefore used as an epileptogenic agent in experimental epileptology. Experiments "in vivo" as well as "in vitro" revealed its relation to NMDA glutamate receptors, and its potential neurotoxicity. From the clinical aspect, hyperhomocysteinemia (HHcy), mostly as a marker of the risk factor in the vascular damage, was often studied in patients treated with antiepileptic drugs (AE). However, the neuroexcitatory influence of mild HHcy (up to 30 micromol/l) was rarely discussed. Out of a group of 123 adult patients on long-term conventional AE we analyzed 8 patients (7 men and one woman) with moderate to severe HHcy (30.7-109.0 micromol/l) retrospectively and 2-5 years after HHcy normalization. All of them suffered from partial and/or secondary generalized seizures accompanied by neuropsychological impairment depending on the aetiology of the disease. The patients were characterized by a concurrence of several factors: (1) All of them received conventional AEs inducing the cytochrome P 450 at the time HHcy was diagnosed. (2) Molecular-genetic tests showed enzymopathic impairment (methylentetrahydrofolate reductase-MTHFR mutation of the gene C677 T) also in all eight, homozygous in 7 cases and heterozygous in 1 case. (3) All patients were found to have a vitamin deficit or marginal values of at least one of the vitamins under study, especially folate and/or vitamin B6 and 812. With reference to clinical and EEG features, the potential neuroexcitatory influence of Hcy is discussed taking into account its effect on pathogenetic factors.

Adult↗

PubMed2004

Phase III study of all-trans retinoic acid in previously untreated patients 61 years or older with acute myeloid leukemia.

The purpose of our study was (i) to evaluate the impact of all-trans retinoic acid (ATRA) given as adjunct to chemotherapy and (ii) to compare second consolidation vs maintenance therapy in elderly patients with acute myeloid leukemia (AML). A total of 242 patients aged >or=61 years (median, 66.6 years) with AML were randomly assigned to ATRA beginning on day +3 after the initiation of chemotherapy (ATRA-arm, n=122) or no ATRA (standard-arm, n=120) in combination with induction and first consolidation therapy. A total of 61 patients in complete remission (CR) were randomly assigned to second intense consolidation (n=31) or 1-year oral maintenance therapy (n=30). After induction therapy the intention-to-treat analysis revealed a significant difference in CR rates between the ATRA- and the standard-arm (52 vs 39%; P=0.05). Event-free (EFS) and overall survival (OS) were significantly better in the ATRA-compared to the standard-arm (P=0.03 and 0.01, respectively). OS after second randomization was significantly better for patients assigned to intensive consolidation therapy (P<0.001). The multivariate model for survival revealed lactate dehydrogenase, cytogenetic risk group, age, and first and second randomization as prognostic variables. In conclusion, the addition of ATRA to induction and consolidation therapy may improve CR rate, EFS and OS in elderly patients with AML.

Acute Disease↗

PubMed2003

Risk-adapted postremission therapy in acute myeloid leukemia: results of the German multicenter AML HD93 treatment trial.

The objective of the AML HD93 treatment trial was to evaluate the outcome in young adults with acute myeloid leukemia (AML) after postremission therapy was stratified according to cytogenetically defined risk. The rationales for the study design were based (i) on previous favorable results with high-dose cytarabine in AML with t(8;21), inv/t(16q22) and in AML with normal karyotype, and ii) on encouraging results obtained in several phase II trials using autologous stem cell transplantation (SCT). Between July 1993 and January 1998, 223 eligible patients, 16-60 years of age with newly diagnosed AML other than French-American-British type M3/M3v, were entered into the trial. Risk groups were defined as follows: low risk: t(8;21) or inv/t(16q22); intermediate risk: normal karyotype; high risk: all other chromosomal abnormalities. Following intensive double induction therapy with idarubicin, cytarabine and etoposide, all patients in complete remission (CR) received a first consolidation therapy with high-dose cytarabine and mitoxantrone (HAM). A second consolidation therapy was stratified according to the risk group: low risk: HAM; intermediate risk: related allogeneic SCT or sequential HAM; high risk: related allogeneic or autologous SCT. Double induction therapy resulted in a high CR rate of 74.5%, and 90% of the responding patients were eligible for consolidation therapy. Survival for all 223 trial entrants was 40%, and for the 166 patients who entered CR, disease-free (DFS) and overall survival were 40 and 51% after 5 years, respectively. Within the low-, intermediate- and high-risk groups, DFS and survival after 5 years were 62.5 and 87, 40 and 49 and 17 and 26% respectively, without advantage for allogeneic transplantation in the intermediate- and high-risk groups. Postremission therapy-related mortality was 0, 7 and 14%, respectively. This study demonstrates the feasibility of cytogenetically defined risk-adapted consolidation therapy. The overall trial results are at least equivalent to those of published trials supporting the risk-adapted treatment strategy.

Acute Disease↗

PubMed2002

Mercury intoxication from skin ointment containing mercuric ammonium chloride.

OBJECTIVE: A one-year follow-up was performed of a 21-year-old man with a 16-year history of diabetes mellitus type I, who had been using ointment containing 10% mercuric ammonium chloride (hydrargyrum amidochloratum; HgNH(2)Cl) for eczema for approximately 3 weeks. Tiredness, fasciculations on the extremities and poor control of diabetes appeared after the end of the ointment treatment. Nephrotic syndrome and hypertension were diagnosed 1 month later. Two months after the ointment application the patient was very weak with tremors of the hands, almost unable to walk, and had lost 20 kg of body weight. He had severe neurasthenic symptoms and his behaviour suggested acute psychosis. METHODS: Internal, neurological and neuropsychological examinations were performed. Mercury in urine was determined by flameless atomic absorption spectrometry. RESULTS: The urine mercury level on admission was 252.0 microg/l. He was treated with Dimaval, sodium (2,3)-dimercaptopropane(-1)-sulphonate capsules for 12 days (total dose 6.3 g). The highest urine mercury excretion during antidote treatment was 2336.0 microg/24 h. The patient had proteinuria of up to 11.10 g/24 h, and renal biopsy revealed diffuse membranous glomerulonephritis of the 1st stage without apparent diabetic nephropathy. Similarly, neuropathy did not have typical signs of diabetic neuropathy. His clinical condition started to improve during the first 2 weeks. Further follow-up has shown slow normalisation of renal functions. After 1 year, proteinuria decreased to 0.62 g/24 h and body weight normalised. Neuropsychological and electromyographic findings became almost normal. CONCLUSION: Severe intoxication developed after a short period of ointment application. Most signs of damage disappeared in the course of 1 year, except mild proteinuria and neuropathy. The evolution was favourable and confirmed the primary role of mercury intoxication in the severe deterioration of the clinical status of the patient.

Administration, Topical↗

PubMed2002

Lipid metabolism and neuropsychological follow-up study of workers exposed to 2,3,7,8- tetrachlordibenzo- p-dioxin.

OBJECTIVE: More than 35 years ago, during 1965-1968, in the former Czechoslovakia, approximately 80 persons became ill due to occupational exposure to 2,3,7,8-tetrachlordibenzo- p-dioxin (2,3,7,8-TCDD). The objective of this study was to investigate the incidence of disorders related to occupational exposure to 2,3,7,8-TCDD. METHODS: Most subjects in the group of 12 former 2,3,7,8-TCDD workers (mean age 56.8 years, exposure 10 days to 23 months) still suffer from disturbances of lipid metabolism, psychic disorders, chloracne, and/or nervous system lesions. All workers were given internal, neuropsychological and ophthalmological examinations. Blood cholesterol and triglycerides were measured, and the common carotid artery was examined by ultrasound (B-mode), with the intima-media thickness (IMT) also being measured. Findings were compared with the 2,3,7,8-TCDD level in 1996. RESULTS: Nine of the 12 previously exposed workers had elevated plasma lipids, and hyperlipidaemia was statistically more frequent in patients with higher 2,3,7,8-TCDD levels ( P=0.03). Subject 1, with the highest 2,3,7,8-TCDD plasma level, had 80% stenosis of the diameter of the carotid artery, which needed acute surgery. Besides him, seven persons had atherosclerotic plaques in the carotid arteries. The mean IMT in the group was 0.85 mm (SD+/-0.19); the normal value is 0.62 mm. Eight subjects had degenerative changes of the ocular fundus. Chloracne was still present in two persons. Neuropsychological findings were assessed as normal only in three persons with lower 2,3,7,8-TCDD plasma levels in 1996. Mean 2,3,7,8-TCDD plasma level in 1996 was 256 pg g(-1) fat (range 14-760). CONCLUSION: Hyperlipidaemia, atherosclerotic plaques, increased IMT, ischaemic heart disease and neuropsychological disturbances were frequent in this group of former 2,3,7,8-TCDD workers. Hyperlipidaemia might have played an important role in most of these disorders. The level of 2,3,7,8-TCDD correlated with the highest level of triglycerides ( P=0.02) and cholesterol ( P=0.01) that was found during the 35-year follow-up. This group belongs to the most heavily 2,3,7,8-TCDD-exposed groups of workers, because the mean estimated concentration at the time of intoxication was approximately 5000 pg g(-1) plasma fat.

Arteriosclerosis↗

PubMed2002

Ectoprotein kinase-mediated phosphorylation of FAT/CD36 regulates palmitate uptake by human platelets.

Glycoprotein IV (FAT/CD36) has been shown to be phosphorylated by a cAMP-dependent, platelet membrane-bound ectokinase. In this study, we demonstrate that ectophosphorylation of FAT/CD36 regulates initial palmitate uptake. This is the first time that short-term regulation of the activity of a long-chain fatty acid carrier could be shown. Phosphorylation of FAT/CD36 was paralleled by a significant decrease in initial palmitate uptake by morphologically and functionally intact platelets. Maximum inhibition of palmitate uptake was achieved at 0.5 nM extracellular ATP, being significantly decreased to 72% compared to the control. Inhibition of palmitate uptake was abolished by co-incubation with the specific protein kinase A inhibitor peptide PKI or with beta,gamma-methylene-ATP, and was reversible upon addition of alkaline phosphatase. An extracellular ATP concentration above 5 microM completely prevented the ectophosphorylation-mediated inhibition of palmitate uptake. We conclude that FAT/CD36-mediated palmitate uptake by human platelets is short-term regulated via cAMP-dependent ectophosphorylation of FAT/CD36.

Blood Platelets↗

PubMed2001

Aspartate residue 142 is important for catalysis by ADP-glucose pyrophosphorylase from Escherichia coli.

Structural prediction of several bacterial and plant ADP-glucose pyrophosphorylases, as well as of other sugar-nucleotide pyrophosphorylases, was used for comparison with the three-dimensional structures of two crystallized pyrophosphorylases (Brown, K., Pompeo, F., Dixon, S., Mengin-Lecreulx, D., Cambillau, C., and Bourne, Y. (1999) EMBO J. 18, 4096-4107; Blankenfeldt, W., Asuncion, M., Lam, J. S., and Naismith, J. H. (2000) EMBO J. 19, 6652-6663). This comparison led to the discovery of highly conserved residues throughout the superfamily of pyrophosphorylases despite the low overall homology. One of those residues, Asp(142) in the ADP-glucose pyrophosphorylase from Escherichia coli, was predicted to be near the substrate site. To elucidate the function that Asp(142) might play in the E. coli ADP-glucose pyrophosphorylase, aspartate was replaced by alanine, asparagine, or glutamate using site-directed mutagenesis. Kinetic analysis in the direction of synthesis or pyrophosphorolysis of the purified mutants showed a decrease in specific activity of up to 4 orders of magnitude. Comparison of other kinetic parameters, i.e. the apparent affinities for substrates and allosteric effectors, showed no significant changes, excluding this residue from the specific role of ligand binding. Only the D142E mutant exhibited altered K(m) values but none as pronounced as the decrease in specific activity. These results show that residue Asp(142) is important in the catalysis of the ADP-glucose pyrophosphorylase from E. coli.

Amino Acid Sequence↗

PubMed2001

Truncated forms of the recombinant Escherichia coli ADP-glucose pyrophosphorylase: the importance of the N-terminal region for allosteric activation and inhibition.

Truncated forms of Escherichia coli ADPglucose pyrophosphorylase were constructed using recombinant DNA techniques. A truncated form of the enzyme having the first 11 amino acid residues from the N-terminus and 2 amino acid residues from the C-terminus deleted was found to be highly active in absence of activator. A 1.6-fold activation by 1.5 mM fructose 1,6 bis-phosphate was observed for the truncated enzyme as compared to the 30-fold activation seen for the intact enzyme. Inhibition of the truncated enzyme by AMP was less than that seen with the intact enzyme. Similar properties were displayed by an enzyme truncated only at the N-terminal. Conversely, the C-terminal truncated enzyme shortened by 2 amino acid residues at the C-terminus is as sensitive as the intact enzyme to activation and inhibition. These results suggest that the N-terminal region is required for allosteric regulation of the enzyme.

Adenosine Monophosphate↗

PubMed2001

Analysis of the amino terminus of maize branching enzyme II by polymerase chain reaction random mutagenesis.

Maize endosperm branching enzyme II (mBEII) plays a pivotal role in determining the quality of starch by catalyzing the synthesis of the alpha-1,6-branch points. While the central (alpha/beta)8-barrel and the C-terminal domains of mBEII have been analyzed previously, the possible role of its amino terminus in catalysis is still poorly understood. Because the amino terminus of mBEII shares very little sequence homology with other amylolytic enzymes, the Met1-Gly276 region of mBEII was randomly mutagenized under error-prone PCR conditions. Subsequent screening by a heterologous complementation system, utilizing an Escherichia coli strain devoid of the endogenous glycogen branching enzyme (glgB-), led to the recovery of mBEII mutants with altered iodine-staining patterns and reduced branching enzyme activities. The NR-625 mutant enzyme, which lacks the N-terminal 39 residues of mBEII due to a frameshift mutation introduced during the random mutagenesis, retained more than 70% of the wild-type activity. The chain transfer pattern and substrate preference of the truncated enzyme were almost identical to those of the wild-type mBEII. It appears that the N-terminal 39 residues of mBEII are neither required for catalysis nor involved in chain transfer. On the other hand, the Gln-to-Arg substitution at position 270 of mBEII resulted in the loss of more than 90% of branching activity. The Gln270 of mBEII, located at the beginning of the (alpha/beta)8-barrel domain, may be required for maximum enzyme activity.

1,4-alpha-Glucan Branching Enzyme↗

PubMed2001

Tyrosine residue 300 is important for activity and stability of branching enzyme from Escherichia coli.

Branching enzyme belongs to the alpha-amylase family, which includes enzymes that catalyze hydrolysis or transglycosylation at alpha-(1,4)- or alpha-(1,6)-glucosidic linkages. In the alpha-amylase family, four highly conserved regions are proposed to make up the active site. From amino acid sequence analysis a tyrosine residue is completely conserved in the alpha-amylase family. In Escherichia coli branching enzyme, this residue (Y300) is located prior to the conserved region 1. Site-directed mutagenesis of the Y300 residue in E. coli branching enzyme was used in order to study its possible function in branching enzymes. Replacement of Y300 with Ala, Asp, Leu, Ser, and Trp resulted in mutant enzymes with less than 1% of wild-type activity. A Y300F substitution retained 25% of wild-type activity. Kinetic analysis of Y300F showed no effect on the Km value. The heat stability of Y300F was analyzed, and this was lowered significantly compared to that of the wild-type enzyme. Y300F also showed lower relative activity at elevated temperatures compared to wild-type. Thus, these results show that Tyr residue 300 in E. coli branching enzyme is important for activity and thermostability of the enzyme.

1,4-alpha-Glucan Branching Enzyme↗

PubMed2001

Starchless mutants of Chlamydomonas reinhardtii lack the small subunit of a heterotetrameric ADP-glucose pyrophosphorylase.

ADP-glucose synthesis through ADP-glucose pyrophosphorylase defines the major rate-controlling step of storage polysaccharide synthesis in both bacteria and plants. We have isolated mutant strains defective in the STA6 locus of the monocellular green alga Chlamydomonas reinhardtii that fail to accumulate starch and lack ADP-glucose pyrophosphorylase activity. We show that this locus encodes a 514-amino-acid polypeptide corresponding to a mature 50-kDa protein with homology to vascular plant ADP-glucose pyrophosphorylase small-subunit sequences. This gene segregates independently from the previously characterized STA1 locus that encodes the large 53-kDa subunit of the same heterotetramer enzyme. Because STA1 locus mutants have retained an AGPase but exhibit lower sensitivity to 3-phosphoglyceric acid activation, we suggest that the small and large subunits of the enzyme define, respectively, the catalytic and regulatory subunits of AGPase in unicellular green algae. We provide preliminary evidence that both the small-subunit mRNA abundance and enzyme activity, and therefore also starch metabolism, may be controlled by the circadian clock.

Amino Acid Sequence↗

PubMed2000

Studies on the effect of temperature on the activity and stability of cyanobacterial ADP-glucose pyrophosphorylase.

The effect of temperature on the activity and stability of ADPglucose pyrophosphorylase from Anabaena PCC 7120 was studied. Experimental optima temperatures were found around 37-40 degrees C or 42-45 degrees C, depending on the absence or the presence of allosteric effectors in the assay medium, respectively. In the range of temperature where the enzyme is stable, curved Arrhenius plots were obtained, indicating a transition temperature between 9 and 12 degrees C. Since these results were observed for both the forward and reverse reaction, with two different sets of substrates and two entirely different assay procedures, it seems unlikely that the effect can be on any component of the system other than the enzyme itself. Results suggest that cyanobacterial ADPglucose pyrophosphorylase undergoes conformational changes at different temperatures, rendering structures with different catalytic efficiencies. The different structures of the enzyme were visualized by emission fluorescence. ADPglucose pyrophosphorylase was irreversibly inactivated when exposed to temperatures above 40 degrees C. Inactivation was dependent on temperature and followed first order kinetics. The substrate, ATP, and the allosteric effectors, 3PGA and Pi, effectively protected the enzyme against thermal inactivation. Protection afforded by ATP was affected by MgCl2. These results suggest that the binding of the effectors to the enzyme resulted in conformational changes of the protein, rendering structures more stable to temperature treatments. Similar structures could be adopted by the enzyme in different environments, since the higher stability was observed in media containing either high ionic strength or high hydrophobicity.

Adenosine Diphosphate Glucose↗

PubMed2000

Localization of C-terminal domains required for the maximal activity or for determination of substrate preference of maize branching enzymes.

Previous analysis of a chimeric enzyme mBEII-IBspHI, in which the C-terminal 229 amino acids of maize endosperm branching enzyme isoform II (mBEII) are replaced by the corresponding 284 amino acids of isoform I (mBEI), suggested that the carboxyl terminus of maize branching enzymes may be involved in catalytic efficiency and substrate preference. In the present study, additional hybrids of mBEI and mBEII were generated and expressed in Escherichia coli BL21 (DE3) to dissect the structure/function relationships of the C-terminal regions of maize branching enzymes. A truncated form of purified mBEII-IBspHI, which lacks the C-terminal 58 amino acids, retained similar levels of V(max) in branching activity, K(m) for reduced amylose AS 320, and substrate preference for amylose than amylopectin when compared to mBEII-IBspHI. This indicates that the C-terminal extension derived from mBEI is not required for either catalysis or substrate preference. However, deletion of an additional 87 amino acids from the carboxyl terminus resulted in complete loss of activity. Replacement of the deleted C-terminal additional 87 amino acids with the corresponding 79 amino acids from mBEII restored 25% of the mBEII-IBspHI branching activity without altering substrate preference. It thus appears that a C-terminal region encompassing Leu649-Asp735 of mBEII-IBspHI is required for maximum catalytic efficiency. Another C-terminal region, residues Gln510-Asp648, of mBEII-IBspHI (Gln476-Asp614 of mBEI) may be involved in substrate-preference determination.

1,4-alpha-Glucan Branching Enzyme↗

PubMed2000

Limited proteolysis of branching enzyme from Escherichia coli.

Branching enzyme is involved in determining the structure of starch and glycogen. It catalyzes the formation of branch points by cleavage and transfer of alpha-1,4-glucan chains to alpha-1,6 branch points. Branching enzyme belongs to the amylolytic family of enzymes containing four conserved regions in a central (alpha/beta)8-barrel. Limited proteolysis of the branching enzyme from Escherichia coli (84 kDa) by proteinase K produced a truncated protein of 70-kDa, which still retained 40-60% of branching activity, depending on the type of assay used. Amino acid sequencing showed that the 70-kDa protein lacked 111 or 113 residues at the amino terminal, whereas the carboxy terminal was still intact. We purified this truncated enzyme to homogeneity and analyzed its properties. The enzyme had a three- to fourfold lower catalytic efficiency than the native enzyme, whereas the substrate specificity was unaltered. Furthermore, a branching enzyme with 112 residues deleted at the amino terminal was constructed by recombinant technology and found to have properties identical to those of the proteolyzed enzyme.

1,4-alpha-Glucan Branching Enzyme↗

PubMed2000

Slow-binding inhibition of branching enzyme by the pseudooligosaccharide BAY e4609.

Branching enzyme from Escherichia coli is shown to be inhibited by the pseudooligosaccharide BAY e4609. The mechanism of binding is studied in detail by kinetics using reduced amylose as substrate. Lineweaver-Burk plots suggest the mechanism of a noncompetitive or slow-binding inhibitor. Further studies by progress curves and rate of loss of branching activity allows us to conclude BAY e4609 as being a slow-binding inhibitor of branching enzyme. We discuss how these results parallel the inhibition of alpha-amylase by acarbose and the significance of branching enzyme as belonging to the amylolytic family.

1,4-alpha-Glucan Branching Enzyme↗

PubMed2000

Activation of the potato tuber ADP-glucose pyrophosphorylase by thioredoxin.

The potato tuber (Solanum tuberosum L.) ADP-glucose pyrophosphorylase (ADP-GlcPPase) catalyzes the first committed step in starch biosynthesis. The main type of regulation of this enzyme is allosteric, and its activity is controlled by the ratio of activator, 3-phosphoglycerate to inhibitor, P(i). It was reported (Fu, Y., Ballicora, M. A., Leykam, J. F., and Preiss, J. (1998) J. Biol. Chem. 273, 25045-25052) that the enzyme was activated by reduction of the Cys(12) disulfide linkage present in the catalytic subunits. In this study, both reduced thioredoxin f and m from spinach (Spinacia oleracea) leaves reduced and activated the enzyme at low concentrations (10 microM) of activator (3-phosphoglycerate). Fifty percent activation was at 4.5 and 8.7 microM for reduced thioredoxin f and m, respectively, and 2 orders of magnitude lower than for dithiothreitol. The activation was reversed by oxidized thioredoxin. Cys(12) is conserved in the ADP-GlcPPases from plant leaves and other tissues except for the monocot endosperm enzymes. We postulate that in photosynthetic tissues, reduction could play a role in the fine regulation of the ADP-GlcPPase mediated by the ferredoxin-thioredoxin system. This is the first time that a covalent mechanism of regulation is postulated in the synthesis of starch.

Allosteric Regulation↗

PubMed2000

Crystallization and preliminary X-ray diffraction analysis of the catalytic subunit of ADP-glucose pyrophosphorylase from potato tuber.

ADP-glucose pyrophosphorylase is the key regulatory enzyme in the biosynthesis of starch in plants and glycogen in bacteria. The enzyme from potato tuber is comprised of a regulatory subunit and a catalytic subunit and is present as a heterotetramer (alpha(2)beta(2)). The catalytic subunit from potato tuber (50 kDa) was crystallized in four different forms, two of which are suitable for structural studies. A tetragonal crystal form obtained in the presence of the substrate analog Cr-ATP diffracted to 2.2 A and belongs to space group P4(1) (or its enantiomorph), with unit-cell parameters a = b = 110.57, c = 190.14 A. A second crystal form obtained diffracted to 2.8 A and belongs to space group P2, with unit-cell parameters a = 80.06, b = 138.84, c = 92.20 A, beta = 112. 40 degrees. As this protein displays no significant homology to any currently known protein structure, a search for heavy-atom derivatives has been initiated.

Adenosine Triphosphatases↗