Biomedical subjects

B Berger

Publications and source records attributed to B Berger.

At least 37 records · Page 2Linked to original sources

PubMed2001

Spectroscopic characterization of metallothionein from the terrestrial snail, Helix pomatia.

The Cd-sequestering metallothionein (MT) isoform isolated from the midgut gland of Roman snails exposed to Cd supplements in the feed was characterized by compositional and spectroscopic analysis. The preparations contained nearly 5 mol of Cd, small amounts of Cu and about 1 mol of Zn per chain mass of 6620 Da, in numerical agreement with the apoprotein's measured capacity of firmly binding a maximum of 6 equivalents of Cd per molecule. As with other Cd-containing MTs the occurrence of a prominent Cd-mercaptide-specific shoulder at 250 nm in its absorption spectrum showed that Cd is complexed in tetrahedral symmetry by the cysteine residues of the protein, and the multiphasic ellipticity profile in the CD spectrum revealed that these complexes are joined to form one or more oligonuclear Cd-mercapto clusters. Both spectral features vanished with the removal of the metal but were reconstituted to maximum amplitudes by readdition of Cd to the metal-free apoprotein, provided precautions were taken to prevent air oxidation of the latter. Quantitative analysis of snail MT reconstituted with Cd established that the 18 cysteine side chains bind the metal in a 3-to-1 ratio; spectroscopic studies on fractionally restored forms demonstrated that the six Cd ions were bound to the apoprotein molecule in succession in two sets of three Cd ions each. Thus, one can infer from the observed stoichiometry and the coordinating preferences of Cd that this gastropod MT, like the Cd-bearing MTs of marine crustaceans, harboured the metal in two separate cyclically constructed Cd3Cys9 clusters. The snail clusters differed, however, from other MTs in their response to acidification. Their protolytic dissociation proceeded through two separate protonation steps with the manifestation of spectroscopically distinguishable intermediate forms. Thus, this snail isoform displays in its metal composition and its chemical and spectroscopic features both similarities and differences to other animal kingdom MTs. Its properties suggest that it serves an important role in the protection of the terrestrial gastropod from Cd.

Animals↗

PubMed2001

[Opisthorchiasis simulating a malignancy].

We report on 2 patients from Siberia suffering from an infection with the parasite Opisthorchis felineus. The unusual course of their disease pretended in case 1 an eosinophilic leukemia and in case 2 a malignoma of the gallbladder. The Opisthorchiasis is endemic in large areas of Asia and Russia. Humans acquire the infection by eating raw fresh-water fish. Symptoms are nonspecific, but detection of eosinophilia in travellers or residents of endemic areas should induce analysis for specific antibodies against Opisthorchis species. Opisthorchiasis is known to be a precursor of cholangiocarcinoma. Malignoma which was initially suspected could be excluded in both cases and the patients were cured by oral administration of Praziquantel, 40-75 mg/kg body weight for 1 day in 3 divided doses.

Adult↗

PubMed2001

When autosomal short tandem repeats fail: optimized primer and reaction design for Y-chromosome short tandem repeat analysis in forensic casework.

Y-chromosomal short tandem repeats (Y-STRs) are useful forensic DNA markers in investigation of sexual assault cases when a mixture of male and female DNA (e.g., in vaginal swabs) is present in a sample, especially when DNA of the male contributor is present only in very small amount compared to the DNA of the female victim. With autosomal STR analysis of male and female DNA, male DNA in mixtures can usually be detected and correctly interpreted only when it exceeds 5%. However, the amplification of some Y-STRs is known to result in polymerase chain reaction (PCR) products that are not associated with the Y-chromosome, but derive from the X-chromosome and/or autosomal regions. This can cause problems in the interpretation of results, particularly when female DNA is present in excess. Consequently, more specific and sensitive Y-STR primers and PCR conditions are needed. This paper presents two casework examples in which sensitive Y-STR multiplexes (with the addition of PCR enhancer) were successfully used in the analysis of mixtures of male and female DNA, the male component not interpretable by standard autosomal STR typing.

Adolescent↗

PubMed2000

Cryopreservation of spermatozoa in cyprinid fishes.

The present study investigated semen cryopreservation in cyprinid fish using computer-assisted sperm motility analysis for viability control. Spermatozoa of the bleak, Chalcalbumus chalcoides, were used as a basic model to describe the toxic and cryoprotective effects of internal and external cryoprotectants, their most effective concentrations and combinations, the freezing and thawing conditions, and the effects of equilibration. We also used these data to develop a cryopreservation protocol for Barbus barbus, Chondrostoma nasus, Ctenopharyngodon idella, Cyprinus cario, Hypohtalmichthys molitrix, Leuciscus cephalus, Rutilus meidingerii, and Vimba vimba. For all investigated species the optimal extender composition was a buffered physiological sperm motility-inhibiting saline solution containing 10% DMSO and 0.5% glycin. The optimal sperm equilibration period in the extender was < or = 5 min. Freezing was performed in an insulated box in liquid nitrogen vapor and it was optimal at 4 to 5 cm above the surface of the liquid, depending on the species. Thawing was optimal in a 25 degrees C water bath whereby the thawing time ranged depending on species from 15 to 45 sec. This cryopreservation protocol resulted in frozen-thawed semen with 35 to 65% motile and 5 to 25% locally motile spermatozoa depending on the quality of fresh semen.

Animals↗

PubMed2000

"Local rules" theory applied to polyomavirus polymorphic capsid assemblies.

The papovaviruses are nonenveloped dsDNA viruses whose capsids are characterized by a non-quasi-equivalent bonding pattern in which 72 pentameric capsomeres occupy positions having either five or six neighboring capsomeres. The local rules theory of Berger et al. (1994, Proc. Natl. Acad. Sci. USA, 91, 7732-7736), previously developed to explain aspects of icosahedral capsid assembly, has been applied to the papovavirus geometry. Local rules describe capsid symmetry patterns in terms of the local interactions of assembly units, such as coat proteins or capsomeres. Polymorphic assemblies, including T = 1 icosahedral, dodecahedral, spiral, and tubular structures of the polyomavirus VP1 protein, can be induced by specific mutations or changes in the solvent conditions during in vitro assembly of the recombinant coat protein. Local rules models were developed to model the wild-type capsid and several polymorphic assemblies. Some assemblies corresponded to structures modeled by small deviations from wild-type local rules. We conclude that aspects of polyomavirus assembly are consistent with local rules models, although they do not explain all polymorphisms. These results may provide insights into the nature of papovavirus assembly, constraints on assembly pathways, and strategies for disrupting assembly.

Capsid↗

PubMed2000

Prisoners of liberation: a psychoanalytic perspective on disenchantment and burnout among career women lawyers.

Using a psychoanalytic perspective, this article addresses the roots and treatment of disillusionment and incipient burnout in female corporate lawyers. It suggests that one of the primary issues that needs to be addressed in therapy with this group is the tendency to be self-punishing, a characteristic that often may be traced back to insensitive parenting. This formulation, combined with the penchant of law firms to regard as "good and expected" the inclination of their workers (especially women) to work without respite and with little regard for their own needs, places individuals at high risk for burnout. A case example is used to illustrate this phenomenon. It is concluded that whereas psychoanalytic treatment greatly may help overstressed professionals, society at large must address the values that foster the attitude of high career achievement at any cost.

Adult↗

PubMed2000

Concordance of the toxicity of pharmaceuticals in humans and in animals.

This report summarizes the results of a multinational pharmaceutical company survey and the outcome of an International Life Sciences Institute (ILSI) Workshop (April 1999), which served to better understand concordance of the toxicity of pharmaceuticals observed in humans with that observed in experimental animals. The Workshop included representatives from academia, the multinational pharmaceutical industry, and international regulatory scientists. The main aim of this project was to examine the strengths and weaknesses of animal studies to predict human toxicity (HT). The database was developed from a survey which covered only those compounds where HTs were identified during clinical development of new pharmaceuticals, determining whether animal toxicity studies identified concordant target organ toxicities in humans. Data collected included codified compounds, therapeutic category, the HT organ system affected, and the species and duration of studies in which the corresponding HT was either first identified or not observed. This survey includes input from 12 pharmaceutical companies with data compiled from 150 compounds with 221 HT events reported. Multiple HTs were reported in 47 cases. The results showed the true positive HT concordance rate of 71% for rodent and nonrodent species, with nonrodents alone being predictive for 63% of HTs and rodents alone for 43%. The highest incidence of overall concordance was seen in hematological, gastrointestinal, and cardiovascular HTs, and the least was seen in cutaneous HT. Where animal models, in one or more species, identified concordant HT, 94% were first observed in studies of 1 month or less in duration. These survey results support the value of in vivo toxicology studies to predict for many significant HTs associated with pharmaceuticals and have helped to identify HT categories that may benefit from improved methods.

Animals↗

PubMed2000

Isolation and characterization of a self-sufficient one-domain protein. (Cd)-metallothionein from Eisenia foetida.

Earthworms have been shown to accumulate trace elements in general, and particularly high amounts of metal ions such as cadmium, copper and zinc. The earthworm's response to metal contamination has been linked to the induction and expression of metallothionein (MT) proteins, a detoxification strategy analogous to that found in other biological systems. The present study focuses on an inducible Cd-MT isolated from the compost-dwelling brandling worm Eisenia foetida (Savigny). A full characterization of the protein (including protein induction, MT cDNA, amino-acid sequence and metal stoichiometry) revealed a new dimension of knowledge to the molecular genetic information available to date. Whereas the elucidated cDNA codes for a putative protein which possesses 80 amino-acid residues, the characterized protein bears only 41 amino acids. The isolated product has evidently attained its size and shape by cleavage near the N-terminal site and at the linker region between the two putative metal-binding domains of the translated product, yielding a small MT moiety which contains 12 Cys residues (including one triple Cys-motif) binding four cadmium ions. It can be shown that the isolated MT molecule represents a self-sufficient one-domain MT which is stable in vitro. The isolation of the single-domain MT peptide raises the question about the method of formation and significance in vivo of such small MT moieties from tissues of E. foetida and possibly other terrestrial invertebrates. In this respect, two hypotheses are discussed: firstly, the possibility of formation of small MT peptides due to enzymatic cleavage of the intact protein during the process of preparation and isolation; and secondly, the possibility of deliberate post-translational processing of the translated gene product to yield functional one-domain MT moieties.

Amino Acid Sequence↗

PubMed2000

Random C-peptide in the classification of diabetes.

OBJECTIVE: To clarify whether random C-peptide is a valuable test in the classification of diabetes. RESEARCH DESIGN AND METHODS: All C-peptide measurements conducted in the diabetic population of Skaraborg (280,539 inhabitants and 3.2% diabetes) between 1995 and 1998 (3,115 samples) were considered, but only patients with well-defined diabetes type (1,449 samples from 1,093 patients) were analyzed for the correlation between diabetes type and C-peptide concentration. Serum C-peptide was measured after fasting over night (fCP), after glucagon stimulation (gCP), and randomly (rCP) without considering previous meals at an ordinary visit to the diabetic clinic (rCP). Receiver Operating Characteristic (ROC) curves were constructed to illustrate the power of the different C-peptide protocols and to determine the optimal cut-off values. RESULTS: Although all three tests had high discriminative power, the ROC curves demonstrated that rCP was superior to fCP and gCP in discriminating type 1 from type 2 diabetes. The optimal cut-off value for rCP was 0.50 nmol/L, for fCP 0.42 nmol/L, and for gCP 0.60 nmol/L. CONCLUSIONS: rCP is more powerful than fCP and gCP in distinguishing type 1 from type 2 diabetes and can therefore be recommended as a classification tool, particularly in outpatients.

Adolescent↗

PubMed2000

Linker insertion mutagenesis based on IS21 transposition: isolation of an AMP-insensitive variant of catabolic ornithine carbamoyltransferase from Pseudomonas aeruginosa.

The bacterial insertion sequence IS21 when repeated in tandem efficiently promotes non-replicative cointegrate formation in Escherichia coli. An IS21-IS21 junction region which had been engineered to contain unique SalI and BglII sites close to the IS21 termini was not affected in the ability to form cointegrates with target plasmids. Based on this finding, a novel procedure of random linker insertion mutagenesis was devised. Suicide plasmids containing the engineered junction region (pME5 and pME6) formed cointegrates with target plasmids in an E.coli host strain expressing the IS21 transposition proteins in trans. Cointegrates were resolved in vitro by restriction with SalI or BglII and ligation; thus, insertions of four or 11 codons, respectively, were created in the target DNA, practically at random. The cloned Pseudomonas aeruginosa arcB gene encoding catabolic ornithine carbamoyltransferase was used as a target. Of 20 different four-codon insertions in arcB, 11 inactivated the enzyme. Among the remaining nine insertion mutants which retained enzyme activity, three enzyme variants had reduced affinity for the substrate ornithine and one had lost recognition of the allosteric activator AMP. The linker insertions obtained illustrate the usefulness of the method in the analysis of structure-function relationships of proteins.

Adenosine Monophosphate↗

PubMed2000

Human and mouse gene structure: comparative analysis and application to exon prediction.

We describe a novel analytical approach to gene recognition based on cross-species comparison. We first undertook a comparison of orthologous genomic loci from human and mouse, studying the extent of similarity in the number, size and sequence of exons and introns. We then developed an approach for recognizing genes within such orthologous regions by first aligning the regions using an iterative global alignment system and then identifying genes based on conservation of exonic features at aligned positions in both species. The alignment and gene recognition are performed by new programs called and, respectively. performed well at exact identification of coding exons in 117 orthologous pairs tested.

Amino Acids↗

PubMed2000

Metallothioneins in terrestrial invertebrates: structural aspects, biological significance and implications for their use as biomarkers.

During the last few years the subject of metallothioneins (MTs) in terrestrial invertebrates has gained increasing attention. One reason for this may be that terrestrial invertebrates provide new insights into the biological diversity of MTs, with the potential of discovering alternative models of structural and functional relationships. Four groups of terrestrial invertebrates have been studied in detail, namely nematodes, insects, snails and earthworms, with the present article focusing on MTs from the latter two groups. Snails are interesting because they possess distinct MT isoforms involved in different metal-specific tasks. In the Roman snail (Helix pomatia), for example, one isoform is predominantly expressed in the midgut gland, accounting for the accumulation, binding and detoxification of cadmium. The second isoform, which is present in the snail's mantle, is substantially different regarding its primary structure. Furthermore, it binds nearly exclusively copper, and thus is probably involved in the homeostatic regulation of essential trace elements. Earthworm MTs merit our attention because of another peculiarity: they seem to be much more unstable than snail MTs, particularly under conventional conditions of preparation. The cDNA of the brandling worm (Eisenia foetida), for instance, codes for a putative MT, which is about twice the size of the actual protein. The isolated MT peptide binds four Cd2+ ions and represents a one-domain MT entity that is stable and functional in vitro. This strongly suggests that earthworm MTs are either posttranslationally modified, or subjected to enzymatic cleavage during preparation. Both snail and earthworm MTs are inducible by metal exposure, especially by cadmium, thus supporting the idea of using them as potential biomarkers for environmental metal pollution. Whilst snail MTs have already been tested in this respect with some success, the use of earthworm MTs as biomarkers still remains to be evaluated, especially in the light of the unknown significance of their posttranslational instability.

Amino Acid Sequence↗

PubMed1999

Sperm metabolism of the telost fishes Chalcalburnus chalcoides and Oncorhynchus mykiss and its relation to motility and viability.

In the teleost fish Chalcalburnus chalcoides (Cyprinidae) the influence of metabolic inhibitors, substrates, coenzymes, and oxygen concentrations on spermatozoal parameters during motility and during immotile incubation was studied, the respiration rate was characterized, representative metabolite levels were measured, and the results were compared with Oncorhynchus mykiss (Salmonidae). In Chalcalburnus chalcoides the sperm motility rate, the average path swimming velocity, the motility duration, and the viability of immotile semen were significantly reduced in the presence of inhibitors of respiration (potassium cyanide, 2.4-dinitrophenol, atractyloside). Anaerobic conditions (<1 mg O(2)/liter) and inhibition of the tricarboxylic acid cycle by malonate and >7.5 mmol/liter succinate had similar effects on the sperm motility parameters and on the viability of immotile spermatozoa. Pyruvate and coenzyme A (an acyl-group carrier during oxidative carboxylation of pyruvate) prolonged the duration of sperm motility and the viability of immotile incubated spermatozoa, and also increased the spermatozoal respiration rate. Glucose levels significantly decreased during motility and during immotile storage and, under anaerobic conditions, the levels of lactate increased indicating that pyruvate derived from glycolysis. The respiration rate and the glycolytic rate significantly increased during motility. Therefore oxidative phosphorylation, tricarboxylic acid cycle, and aerobic glycolysis are central energy-supplying pathways for spermatozoa of Chalcalburnus chalcoides. The stimulatory effect of pyruvate and coenzyme A indicated that glycolysis is a rate-controlling pathway. Similar results were obtained for Oncorhynchus mykiss with the only exception that the stimulatory effect of coenzyme A was more significant than the stimulatory effect of pyruvate. When the sperm motility-activating saline solutions were optimized in aspects of energy supply, ionic composition, and osmolality, about 50% of the motile spermatozoa swam progressively (>20 mm/sec) for about 3 min in Chalcalburnus chalcoides and in Oncorhynchus mykiss. About 20% swam progressively for >2 hr in Chalcalburnus chalcoides and for >30 min in Oncorhynchus mykiss. J. Exp. Zool. 284:454-465, 1999.

2,4-Dinitrophenol↗

PubMed1999

LearnCoil-VMF: computational evidence for coiled-coil-like motifs in many viral membrane-fusion proteins.

Crystallographic studies have shown that the coiled-coil motif occurs in several viral membrane-fusion proteins, including HIV-1 gp41 and influenza virus hemagglutinin. Here, the LearnCoil-VMF program was designed as a specialized program for identifying coiled-coil-like regions in viral membrane-fusion proteins. Based upon the use of LearnCoil-VMF, as well as other computational tools, we report detailed sequence analyses of coiled-coil-like regions in retrovirus, paramyxovirus and filovirus membrane-fusion proteins. Additionally, sequence analyses of these proteins outside their putative coiled-coil domains illustrate some structural differences between them. Complementing previous crystallographic studies, the coiled-coil-like regions detected by LearnCoil-VMF provide further evidence that the three-stranded coiled coil is a common motif found in many diverse viral membrane-fusion proteins. The abundance and structural conservation of this motif, even in the absence of sequence homology, suggests that it is critical for viral-cellular membrane fusion. The LearnCoil-VMF program is available at http://web.wi.mit.edu/kim

Amino Acid Sequence↗

PubMed1999

Precocious development of parvalbumin-like immunoreactive interneurons in the hippocampal formation and entorhinal cortex of the fetal cynomolgus monkey.

The calcium-binding protein parvalbumin (PV), a reliable marker of the hippocampal basket and chandelier cells, is first expressed on embryonic day 83 (E83), corresponding to midgestation of the macaque monkey, in restricted hippocampal groups of immature neurons (Berger and Alvarez [1996] J. Comp. Neurol. 366:674-699). In the present study, PV-like immunoreactivity (LIR) was used to follow the further development of this subclass of interneurons. Asynchronous area-specific developmental sequences were observed, predominating initially in the caudal half of the hippocampal formation and the laterocaudal division of the entorhinal cortex and occurring relatively simultaneously in the interconnected hippocampal and entorhinal subfields. Dendritic elongation of PV-like immunoreactive interneurons and perisomatic distribution of PV-like immunoreactive terminal boutons on their cellular targets were first observed in the subiculum around E127; then from E127 to E142 in CA3/CA2 and layers III-V of the entorhinal cortex and, to a lesser extent in CA1, the dentate hilus and deep granule cell layer; and finally from E156 to postnatal day 12 in the rest of the dentate gyrus, the presubiculum and parasubiculum, and layers III-II-I of the entorhinal cortex. These data provide the first indication that a population of basket cells, a major gamma-aminobutyric acid (GABA)ergic component of the hippocampal intrinsic inhibitory circuitry, reaches its cellular targets several weeks before birth in primates in contrast to rodents. The role of the prenatal PV expression in the hippocampal formation of nonhuman primates and whether it coincides with the onset of postsynaptic inhibitory potentials or is accompanied or preceded by a period of gamma-aminobutyric acid-mediated excitatory effects as in rat pups, are crucial questions. They underline the need to pursue direct investigations on primates to be able to legitimately extrapolate the data obtained in rodents.

Aging↗

PubMed1999

On the structure of the scaffolding core of bacteriophage T4.

The scaffolding core in bacteriophages is a temporary structure that plays a major role in determining the shape of the protein shell that encapsulates the viral DNA. In the currently accepted structure for the scaffolding core in bacteriophage T4, there is a symmetry mismatch between the protein shell, which has fivefold symmetry, and the scaffolding core, which is believed to consist of six helical chains. The analysis of T4 giant prohead data that was used to determine this structure made an implicit assumption about the manner in which giant proheads flatten during preparation for electron microscopy. Namely, it was assumed that techniques for analysis of Fourier transforms of flattened single-layer cylinders could be applied independently to the shell and the core. This analysis makes the implicit assumption that connections between the core and the shell do not affect the flattening process, and thus are stretched or broken during the flattening process. Reexamination of the experimental data shows that this assumption is likely to be incorrect. A reanalysis shows that the data could be consistent with six, eight, or 10 helical chains, and is a better match for eight or 10 helical chains. Ten helical chains would match the fivefold symmetry of the shell. The 10-helix core model is particularly attractive because it suggests a Vernier mechanism, which is able to explain the process of length determination in giant head mutants of T4. It is possible that the same assumption has been made for structural analysis of other biological systems. If this is the case, any results obtained should also be reexamined.

Bacteriophage T4↗

PubMed1999

Sequencing a genome by walking with clone-end sequences: a mathematical analysis.

One approach to sequencing a large genome is (1) to sequence a collection of nonoverlapping "seeds" chosen from a genomic library of large-insert clones [such as bacterial artificial chromosomes (BACs)] and then (2) to take successive "walking" steps by selecting and sequencing minimally overlapping clones, using information such as clone-end sequences to identify the overlaps. In this paper we analyze the strategic issues involved in using this approach. We derive formulas showing how two key factors, the initial density of seed clones and the depth of the genomic library used for walking, affect the cost and time of a sequencing project-that is, the amount of redundant sequencing and the number of steps to cover the vast majority of the genome. We also discuss a variant strategy in which a second genomic library with clones having a somewhat smaller insert size is used to close gaps. This approach can dramatically decrease the amount of redundant sequencing, without affecting the rate at which the genome is covered.

Chromosome Walking↗

PubMed1999

Target joining of duplicated insertion sequence IS21 is assisted by IstB protein in vitro.

Tandemly repeated insertion sequence IS21, located on a suicide plasmid, promoted replicon fusion with bacteriophage lambda in vitro in the presence of ATP. This reaction was catalyzed in a cell extract containing the 45-kDa IstA protein (cointegrase) and the 30-kDa IstB helper protein of IS21 after both proteins had been overproduced in Escherichia coli. Without IstB, replicon fusion was inefficient and did not produce the 4-bp target duplications typical of IS21.

Artificial Gene Fusion↗