Biomedical subjects

B Svensson

Publications and source records attributed to B Svensson.

At least 163 records · Page 9Linked to original sources

PubMed1993

Detection of a purine analogue-sensitive kinase in frog sciatic nerves--possible involvement in nerve regeneration.

This study reports the existence of a purine analogue-sensitive protein kinase in adult frog sciatic nerves. Cell-free supernatants of homogenized regenerating sciatic nerves were found to contain a phosphoprotein (MW 90 kDa, referred to as PP90), that was phosphorylated to a much higher degree than in normal, uninjured nerves. The spatial and temporal characteristics of PP90 phosphorylation suggested a relationship with the injury-induced proliferation of support cells of the regenerating nerve, i.e. its appearance and increment over time correlated with that of [3H]thymidine incorporation in the nerve. PP90 was phosphorylated under conditions that excluded enzyme activities due to Ca2+/calmodulin kinases, cyclic nucleotide-dependent kinases or protein kinase C. On the other hand, the phosphorylation could be selectively inhibited by the purine analogues adenosine, 2-aminopurine and 6-thioguanine (6-TG). The latter was the most potent and gave complete inhibition at 50 microM. Addition of histone H1 to the cell-free assay stimulated the phosphorylation of several proteins in both normal and regenerating nerves. The stimulation could be blocked by 6-TG, indicating the presence of a purine-sensitive kinase also in uninjured nerves. Separate experiments showed that in vitro regeneration of the frog sciatic sensory axons, as well as the proliferation of the support cells, was inhibited by 100 microM 6-TG.

Animals↗

PubMed1993

Arginine is essential for the alpha-amylase inhibitory activity of the alpha-amylase/subtilisin inhibitor (BASI) from barley seeds.

Treatment of barley alpha-amylase/subtilisin inhibitor (BASI) with reagents specific for arginine, histidine, methionine and tyrosine residues and amino and carboxyl groups indicates that an arginine residue(s) is essential for its action on the target enzyme barley alpha-amylase 2. Phenylglyoxal modified eight out of 12 arginine residues in BASI. Kinetic analysis shows that the inactivation of BASI follows a pseudo-first-order reaction and is due to reaction with one molecule of phenylglyoxal; the second-order rate constant is determined to be 2.95 M-1.min-1. At pH 8.0, BASI and barley alpha-amylase 2 form an inactive 1:1 complex. The Ki value of this association is 2.2 x 10(-10) M. The alpha-amylase protects four arginine residues and also the alpha-amylase inhibitory activity of BASI against phenylglyoxal. When BASI from the phenylglyoxal-modified target enzyme-inhibitor complex is isolated and subjected to a second treatment with phenylglyoxal, four additional arginine residues are modified, with concomitant loss of the inhibitory activity. These results are discussed in relation to a three-dimensional model of BASI based on the known structure of the corresponding inhibitor from wheat.

Arginine↗

PubMed1993

Modified EPR spectra of the tyrosineD radical in photosystem II in site-directed mutants of Synechocystis sp. PCC 6803: identification of side chains in the immediate vicinity of tyrosineD on the D2 protein.

The oxidizing side of photosystem II contains two redox-active tyrosyl side chains, TyrZ and TyrD, and a cluster of Mn atoms involved in water oxidation. The structural environment of these components is unknown, and with computer-assisted modeling we have created a three-dimensional model for the structures around TyrZ and TyrD [Svensson et al. (1990) EMBO J. 9, 2051-2059]. Both tyrosines are proposed to form hydrogen bonds to nearby histidine residues (for Synechocystis 6803, these are His190 on the D1 and His189 on the D2 proteins). We have tested this proposal by electron paramagnetic resonance (EPR) spectroscopy of TyrDox in mutants of the cyanobacterium Synechocystis 6803 carrying site-directed mutations in the D2 protein. In two mutants, where His189 of the D2 protein is changed to either Tyr or Leu, the normal EPR spectrum from TyrDox is replaced by narrow, structureless radical signals with g-values similar to that of TyrDox (g approximately 2.0050). The new radicals copurify with photosystem II, are dark-stable, destabilized by elevated pH, and light-inducible, and originate from radicals formed by oxidation. These properties are similar to those of normal TyrDox, and we assign the new spectra to TyrDox in an altered environment induced by the point mutation in His189. In a third mutant, where Gln164 of the D2 protein was mutated to Leu, we also observed a modified EPR spectrum from TyrDox. This is also consistent with the model in which this residue is found in the immediate vicinity of TyrDox. Thus the results provide experimental evidence supporting essential aspects of the structural model.

Computer Simulation↗

PubMed1993

Production, purification and characterization of the catalytic domain of glucoamylase from Aspergillus niger.

The catalytic domain of glucoamylases G1 and G2 from Aspergillus niger is produced in vitro in high yield by limited proteolysis using either subtilisin Novo or subtilisin Carlsberg. Purification by affinity chromatography on an acarbose-Sepharose column followed by ion-exchange chromatography on HiLoad Q-Sepharose leads to separation of a number of structurally closely related forms of domain. The cleavage occurs primarily between Val-470 and Ala-471 as indicated by C-terminal sequencing, whereas the N-terminus is intact. Subtilisin Carlsberg, in addition, produces a type of domain which is hydrolysed before Ser-444, an O-glycosylated residue. This leaves the fragment Ser-444-Val-470 disulphide-bonded to the large N-terminal part of the catalytic domain. Subtilisin Novo, in contrast, tends to yield a minor fraction of forms extending approx. 30-40 amino-acid residues beyond Val-470. The thermostability is essentially the same for the single-chain catalytic domain and the original glucoamylases G1 and G2, whereas the catalytic domain cut between Ser-443 and Ser-444 is less thermostable. For both types of domain the kinetic parameters, Km and kcat., for hydrolysis of maltose are very close to the values found for glucoamylases G1 and G2.

Amino Acid Sequence↗

PubMed1993

Binding of Ca2+ to calmodulin and its tryptic fragments: theory and experiment.

The Ca(2+)-binding constant of the protein calmodulin is determined experimentally at different pH and varying salt concentration. By comparison to statistical mechanical simulation results, it is shown that the shift in binding constant upon addition of salt is almost quantitatively due to electrostatic interactions. Specific interactions as well as effects due to structural rearrangements of the protein seem to be less important, indicating a structurally rather conserved protein upon addition of salt and changes in pH. The tryptic fragments of calmodulin also bind calcium with high affinity, and the electrostatic effects seem likewise to dominate the binding constant shifts in these systems. It is also shown that the chemical potential of free calcium ions, which is normally dominated by the salt, is strongly influenced by the highly charged calmodulin molecule. This complicates a detailed comparison at low salt concentration, since it requires very accurate information on the salt and protein concentrations, which normally are not available.

Animals↗

PubMed1993

Functional roles of the invariant aspartic acid 55, tyrosine 306, and aspartic acid 309 in glucoamylase from Aspergillus awamori studied by mutagenesis.

Three mutants, Asp55-Gly, Tyr306-->Phe, and Asp309-->Asn, of Aspergillus awamori glucoamylase (identical to Aspergillus niger glucoamylase) were constructed to elucidate the roles of two conserved regions within fungal glucoamylases. Kinetic studies indicate that both of these regions are closely associated with activity. The Asp55-->Gly mutation decreases the kcat approximately 200 times toward maltose and isomaltose, while KM values remain similar to the wild-type. This localizes Asp55 to subsite 1 of glucoamylase where it affects catalytic activity, but not ground-state binding. The pKa value of the catalytic general acid, Glu179, is 1 pH unit lower in that mutant compared to wild-type enzyme, confirming the proximity of Asp55 to the site of catalysis. Tyr306-->Phe is highly active, but affects binding in subsite 2. It moreover shows enhanced binding in the fourth subsite, suggesting that the conserved region around residue 306 interacts with Trp120, a critical residue that directs conformational changes stabilizing the transition-state structure. Finally, the Asp309-->Asn mutation decreases the kcat for isomaltose hydrolysis around 200-fold, but only 30-fold for maltose. This specific effect on the hydrolysis of the alpha-1,6-linked substrate locates Asp309 to subsite 2. Substitution of Asp309 influences affinities of distant subsites, especially subsite 4, similar to mutations of other carboxylic acid residues situated near subsites 1 and 2.

Aspartic Acid↗

PubMed1993

Starch- and glycogen-debranching and branching enzymes: prediction of structural features of the catalytic (beta/alpha)8-barrel domain and evolutionary relationship to other amylolytic enzymes.

Sequence alignment and structure prediction are used to locate catalytic alpha-amylase-type (beta/alpha)8-barrel domains and the positions of their beta-strands and alpha-helices in isoamylase, pullulanase, neopullulanase, alpha-amylase-pullulanase, dextran glucosidase, branching enzyme, and glycogen branching enzymes--all enzymes involved in hydrolysis or synthesis of alpha-1,6-glucosidic linkages in starch and related polysaccharides. This has allowed identification of the transferase active site of the glycogen debranching enzyme and the locations of beta-->alpha loops making up the active sites of all enzymes studied. Activity and specificity of the enzymes are discussed in terms of conserved amino acid residues and loop variations. An evolutionary distance tree of 47 amylolytic and related enzymes is built on 37 residues representing the four best conserved beta-strands of the barrel. It exhibits clusters of enzymes close in specificity, with the branching and glycogen debranching enzymes being the most distantly related.

Amino Acid Sequence↗

PubMed1993

Benign osteoblastoma associated with an aneurysmal bone cyst of the mandibular ramus and condyle.

This article reviews the clinical behavior, the histologic conditions, and the treatment of a benign osteoblastoma associated with an aneurysmal bone cyst of the mandibular condyle and ramus. A 14-year-old boy, otherwise healthy, was referred for pain and enlargement in his right temporomandibular joint-cheek region. The swelling was firm and tender, a slight facial asymmetry was present, and the mouth-opening capacity was reduced. During a 6-weeks period from the primary examination to surgery the lesion expanded extensively, anesthesia appeared in the right inferior alveolar nerve and a lateral open bite developed on the affected side. Computed tomography displayed a total destruction of the right mandibular condyle and ramus to the level of the mandibular foramen. The lesion was radiolucent without distinct borders to adjacent bone. Surgery revealed a bluish, well-vascularized predominantly cystic tumor. The lesion included the temporomandibular joint disk but without overgrowth to the temporal component. After resection of the lesion, the condyle and the mandibular ramus was reconstructed with the use of an autogenous costochondral graft. The postoperative healing was uncomplicated. At follow-up 2 years after surgery the boy was free of recurrence with normal motor and sensory nerve function.

Adolescent↗

PubMed1993

Early surgical-orthodontic treatment of mandibular hypoplasia in juvenile chronic arthritis.

This study is an intermediate evaluation of early surgical-orthodontic treatment in 7 patients with uni- or bilateral hypoplastic mandibular growth due to juvenile chronic arthritis. Resection of the condylar heads and reconstruction with costochondral grafts was undertaken between 10 and 14 years of age and followed by orthodontic treatment. At the latest follow-up, 2-4.5 years postoperatively, all patients showed mandibular growth. Some even showed a tendency towards progenia. The scheduling of the surgery in relation to completion of growth seemed to be of importance in relation to the magnitude of mandibular growth. At the latest check-up, only one patient had attained skeletal maturation and a more long-term follow-up is needed before this method can be finally evaluated.

Adolescent↗

PubMed1993

Electrospray mass spectrometry characterization of post-translational modifications of barley alpha-amylase 1 produced in yeast.

We have used electrospray mass spectrometry (ESMS) in combination with protein chemistry and genetics to delineate post-translational modifications in yeast of barley alpha-amylase 1 (AMY1), a 45 kD enzyme crucial for production of malt, an important starting material in the manufacture of beer and whisky. In addition to signal peptide processing these modifications are: (1) removal of C-terminal Arg-Ser by Kex1p, (2) glutathionylation of Cys95, (3) O-glycosylation, and (4) additional degradation of the C-terminus.

Base Sequence↗

PubMed1993

Functional roles and subsite locations of Leu177, Trp178 and Asn182 of Aspergillus awamori glucoamylase determined by site-directed mutagenesis.

Fungal glucoamylases contain four conserved regions. One region from the Aspergillus niger enzyme contains three key carboxylic acid residues, the general acid catalytic group, Glu179, along with Asp176 and Glu180. Three site-directed mutations, Leu177-->His, Trp178-->Arg and Asn182-->Ala, were constructed near these acidic groups to reveal the function of other conserved residues in this region. Leu177 and Trp178 are strictly conserved among fungal glucoamylases, while an amide, predominantly Asn, always occurs at position 182. Substitutions of Leu177 or Trp178 cause significant decreases in kcat with the substrates tested. Similar increases in activation energies obtained with Leu177-->His with both alpha-(1,4)- and alpha-(1,6)-linked substrates indicate Leu177 is located in subsite 1. KM values obtained with the Trp178-->Arg mutation increase for an alpha-(1,6)-linked substrate, but not for alpha-(1,4)-linked substrates. Calculated differences in activation energy between substrates indicate Trp178 interacts specifically with subsite 2. The Asn182-->Ala mutation did not change kcat or KM values, indicating that Asn182 is not crucial for activity. These results support a mechanism for glucoamylase catalytic activity consisting of a fast substrate binding step followed by a conformational change at subsite 1 to stabilize the transition state complex.

Amino Acid Sequence↗

PubMed1993

Bacillus subtilis CtaA and CtaB function in haem A biosynthesis.

Haem A, a prosthetic group of many respiratory oxidases, is probably synthesized from haem B (protohaem IX) in a pathway in which haem O is an intermediate. Possible roles of the Bacillus subtilis ctaA and ctaB gene products in haem O and haem A synthesis were studied. Escherichia coli does not contain haem A. The ctaA gene on plasmids in E. coli resulted in haem A accumulation in membranes. The presence of ctaB together with ctaA increased the amount of haem A found in E. coli. Haem O was not detected in wild-type B. subtilis strains. A previously isolated B. subtilis ctaA deletion mutant was found to contain haem B and haem O, but not haem A. B. subtilis ctaB deletion mutants were constructed and found to lack both haem A and haem O. The results with E. coli and B. subtilis strongly suggest that the B. subtilis CtaA protein functions in haem A synthesis. It is tentatively suggested that if functions in the oxygenation/oxidation of the methyl side group of carbon 8 of haem O. B. subtilis CtaB, which is homologous to Saccharomyces cerevisiae COX10 and E. coli CyoE, also has a role in haem A synthesis and seems to be required for both cytochrome a and cytochrome o synthesis.

Alkyl and Aryl Transferases↗

PubMed1993

Inhibition of exercise-induced shortening of bleeding time by fish oil in familial hypercholesterolemia (type IIa).

Fourteen patients suffering from familial hypercholesterolemia (type IIa) participated in a double-blind, placebo-controlled trial that evaluated the effects of fish oil ethyl ester (K-85, 5.7 g/day) or a hydroxymethylglutaryl coenzyme A reductase inhibitor (lovastatin, 40 mg/day) alone or in combination on lipid metabolism and bleeding time at rest and after standardized exercise. Lovastatin treatment reduced total cholesterol (-27%), low density lipoprotein cholesterol (-37%), and triglycerides (-18%), whereas high density lipoprotein cholesterol increased significantly (14%). K-85 affected total (-4%), low density lipoprotein (-9%), and high density lipoprotein (+7%) cholesterol insignificantly, whereas the triglyceride level decreased by 24% (p < 0.001). The combined regimen caused an additive decrease in the triglyceride level (41%), which differed significantly (p < 0.01) from that gained by lovastatin alone. Under basal conditions the bleeding time was not influenced by the different interventions. Standardized exercise shortened the bleeding time by 19% (p < 0.001) and 16% (p < 0.001) before intervention and after lovastatin treatment, respectively. After K-85 alone or in combination with lovastatin, the exercise-induced shortening of the bleeding time was totally inhibited, which may reflect a favorable influence of fish oil on the platelet-vessel wall interaction in these high-risk patients.

Adult↗

PubMed1993

The bleeding time in women: an influence of the sex hormones?

A sex difference in the bleeding time has been reported. To investigate the effects of the different sex hormones, the bleeding time (BT) and hemostatic factors related to it were measured in 209 healthy women: 50 pregnant, 113 menstruating and 46 post-menopausal. The serum levels of estrogens, progesterone and testosterone were distributed as expected for the different groups. There were no significant differences in the BT between any of the groups. No correlations between the BT and female or male sex hormones were found. The findings indicate that although sex hormone mediated effects on platelet/endothelial cell interactions may exist, they do not become evident within the physiological variations in the BT of healthy women.

Adult↗

PubMed1993

Formation of free radicals during phacoemulsification.

During phacoemulsification cavitation bubbles are formed. These bubbles are believed to be one source of damage to corneal endothelium seen after phacoemulsification. Free radicals are induced whenever cavitation bubbles implode. The aim of this study was to confirm the initiation of free radicals by phacoemulsification and to correlate the power of ultrasound in the phacoemulsification process to the amount of free radicals formed, using both in vitro and in vivo techniques. The formation of free radicals was determined by adding luminol to a buffer and measuring the chemoluminescence in vitro and in rabbit eyes (Lumacounter 2080 or a single-photon-counting apparatus) during phacoemulsification. The data obtained show that free radicals are formed during phacoemulsification and that the amount of free radicals correlates with the power of ultrasound. Furthermore, the radical formation could be inhibited by the radical scavengers SOD, Healon and Healon GV. These results were achieved both in vitro in the test tube and in vivo in rabbit eyes. By showing that the addition of SOD to the irrigation buffer during phacoemulsification decreases the corneal endothelial cell damage, we show that free radicals could have a role in postoperative complications seen clinically.

Animals↗

PubMed1993

Correction of implant malalignment by segmental osteotomy: a case report.

This case report demonstrates that the position of malaligned osseointegrated implants can be corrected by a segmental osteotomy procedure with no loss of implants or bone support. The patient was a healthy teenage girl who sustained facial trauma with loss of teeth and alveolar bone. The defect was initially bone grafted. Brånemark implants were simultaneously placed in the graft. Their initial unfavorable positions and inclinations were later corrected by a segmental osteotomy.

Adolescent↗

PubMed1993

Comparative effects of prolonged intake of highly purified fish oils as ethyl ester or triglyceride on lipids, haemostasis and platelet function in normolipaemic men.

Conflicting results have been reported about the absorption of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) either as an ethyl ester (EE) or in a triglyceride (TG) formula. We decided to conduct a randomized double-blind study to compare the effects of EE and TG on plasma fatty acids, platelet function and haemostasis. Thirty-one healthy normolipaemic men were allocated to receive fish oil concentrate either as EE or TG with equal amounts of EPA (2.2 g and 2.2 g, respectively) and DHA (1.2 g and 1.4 g, respectively) or placebo daily for 7 weeks. Total cholesterol and the triglyceride level were not influenced differently by the two compounds. Repeated measurement ANOVA revealed a difference between TG and EE regarding incorporation of arachidonic acid (P = 0.034) and EPA (P = 0.007) into plasma cholesteryl esters. A discrimination not observed within plasma phospholipids. Both formulas had equal inhibitory effects on collagen-induced platelet aggregation and thromboxane B2 (TxB2) production in whole blood. Fibrinogen decreased 16% in EE (P = 0.034) and 12% in the placebo group (P = 0.11), but variance analysis of delta change during intervention did not indicate differences between groups. It is concluded that TG and EE fish oils are well incorporated into plasma lipids and have similarly beneficial influence one platelet function in men.

Adult↗

PubMed1993

Effects of cod liver oil on lipids and platelets in males and females.

Thirty-four healthy non-smoking subjects (20 men and 14 women) were given 25 ml cod liver oil (CLO) daily as dietary supplement for 8 weeks in a controlled cross-over study. Prior to intervention, women had significantly higher serum total (P < 0.05) and high density lipoprotein (HDL) (P < 0.001) cholesterol values, higher platelet counts (P < 0.05) and lower thromboxane B2 (TxB2) generation (P < 0.01) in response to collagen stimulation in platelet-rich plasma (PRP) than men. Before intervention, no gender differences in platelet aggregation were observed. Following CLO intervention, serum triglycerides were reduced by 0.21 +/- 0.09 mmol/l (P < 0.05) in men. LDL cholesterol increased in men by 0.28 +/- 0.12 mmol/l (P < 0.05), while the values in women decreased insignificantly. The response in LDL cholesterol to CLO supplementation differed significantly between genders (P < 0.05). HDL cholesterol was not altered in any group. Mean platelet volume (MPV) increased by approximately 6% in both sexes (P < 0.01). After CLO intake, platelet sensitivity to collagen in PRP decreased in men only (P < 0.01), and the response was significantly different from that in women (P < 0.05). Thromboxane B2 generation following platelet aggregation in PRP was equally decreased by CLO. Our findings indicate sex differences in the response to CLO regarding LDL cholesterol and platelet sensitivity to collagen in PRP.

Adult↗