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Biomedical subjects

K Hammer

Publications and source records attributed to K Hammer.

At least 55 records · Page 3Linked to original sources
PubMed1994

Two different dihydroorotate dehydrogenases in Lactococcus lactis.

The pyrimidine de novo biosynthesis pathway has been characterized for a number of organisms. The general pathway consists of six enzymatic steps. In the characterization of the pyrimidine pathway of Lactococcus lactis, two different pyrD genes encoding dihydroorotate dehydrogenase were isolated. The nucleotide sequences of the two genes, pyrDa and pyrDb, have been determined. One of the deduced amino acid sequences has a high degree of homology to the Saccharomyces cerevisiae dihydroorotate dehydrogenase, and the other resembles the dihydroorotate dehydrogenase from Bacillus subtilis. It is possible to distinguish between the two enzymes in crude extracts by using different electron acceptors. We constructed mutants containing a mutated form of either one or the other or both of the pyrD genes. Only the double mutant is pyrimidine auxotrophic.

Amino Acid Sequence↗
PubMed1994

Isolation and characterization of Bacillus subtilis genomic lacZ fusions induced during partial purine starvation.

Random genomic Bacillus subtilis lacZ fusions were screened in order to identify the possible existence of regulons responding to the stimuli generated by partial purine starvation. A leaky pur mutation (purL8) was isolated and used to generate the partial purine starvation conditions in the host strain used for screening. On the basis of their induction during partial purine starvation, seven genomic lacZ fusions were isolated. None of the fusions map in loci previously reported to contain purine-regulated genes. One fusion maps very close to the citB locus and may very well be a citB fusion. The fusions were divided into two types on the basis of their response to complete starvation for either ATP or GTP or both components at the same time. Except for one, type 2 fusions were induced by specific starvation for ATP and by simultaneous starvation for ATP and GTP, but not by specific GTP starvation in a gua strain or by GTP starvation induced by the addition of decoyinine. Type 1 fusions were equally well induced by all three kinds of purine starvation including GTP starvation induced by decoyinine. Further subdivisions of the fusions were obtained on the basis of their responses to the spo0A gene product. A total of five fusions showed that spo0A affected expression. One class was unable to induce lacZ expression in the absence of the spo0A gene product, whereas the other class had increased lacZ expression during partial purine starvation in a spo0A background.

Adenosine↗
PubMed1994

Cloning and characterization of upp, a gene encoding uracil phosphoribosyltransferase from Lactococcus lactis.

Uracil phosphoribosyltransferase catalyzes the key reaction in the salvage of uracil in many microorganisms. The gene encoding uracil phosphoribosyltransferase (upp) was cloned from Lactococcus lactis subsp. cremoris MG1363 by complementation of an Escherichia coli mutant. The gene was sequenced, and the putative amino acid sequence was deduced. The promoter was mapped by both primer extension and analysis of beta-galactosidase expressed from strains carrying fusion between upp promoter fragments and the lacLM gene. The results showed that the upp gene was expressed from its own promoter. After in vitro construction of an internal deletion, a upp mutant was constructed by a double-crossover event. This implicated the utilization of a plasmid with a thermosensitive origin of replication and a new and easy way to screen for double crossover events in both gram-positive and gram-negative bacterial strains. The phenotype of the uracil phosphoribosyltransferase-deficient strain was established. Surprisingly, the upp strain is resistant only to very low concentrations of 5-fluorouracil. Secondary mutants in thymidine phosphorylase and thymidine kinase were isolated by selection for resistance to high concentrations of 5-fluorouracil.

Amino Acid Sequence↗
PubMed1994

Characterization of the lactococcal temperate phage TP901-1 and its site-specific integration.

The temperate lactococcal phage TP901-1, induced by UV light from Lactococcus lactis subsp. cremoris 901-1, was characterized. The restriction map was found to be circular, and the packaging of TP901-1 DNA was concluded to occur by a headful mechanism. The pac region was localized on the 38.4-kb phage genome. TP901-1 belongs to the class of P335 phages (V. Braun, S. Hertwig, H. Neve, A. Geis, and M. Teuber, J. Gen. Microbiol. 135:2551-2560, 1989). Evidence is presented that the phages TP936-1 (V. Braun, S. Hertwig, H. Neve, A. Geis, and M. Teuber, J. Gen. Microbiol. 135:2551-2560, 1989) and C3-T1 (A. W. Jarvis, V. R. Parker, and M. B. Bianchin, Can. J. Microbiol. 38:398-404, 1992) are very closely related to or are identical to TP901-1. The lytically propagated TP901-1 phages were able to lysogenize both indicator strains Lactococcus cremoris 3107 and Wg2. Lysogenization resulted in site-specific integration of the phage genome into the bacterial chromosome. Only one chromosomal attB site was found in 20 independent lysogens. The attP region of TP901-1 and the attL and attR regions were cloned and sequenced. The results showed a core region of only 5 bp, in which the recombination occurs, followed after a 1-bp mismatch by a 7-bp identical region, TCAAT(T/C)AAGGTAA. This result was further verified by sequencing of the attB region obtained by PCR. An integration vector was constructed with the 6.5-kb EcoRI fragment from TP901-1 containing attP. This vector also functions in the plasmid-free strains, MG1363 and LM0230 with only one specific attB site, strongly indicating a more general use of the TP901-1-based integration vector in lactococci.

Bacteriophages↗
PubMed1994

Nucleotide metabolism in Lactococcus lactis: salvage pathways of exogenous pyrimidines.

By measuring enzyme activities in crude extracts and studying the effect of toxic analogs (5-fluoropyrimidines) on cell growth, the metabolism of pyrimidines in Lactococcus lactis was analyzed. Pathways by which uracil, uridine, deoxyuridine, cytidine, and deoxycytidine are metabolized in L. lactis were established. They are similar to those found in Escherichia coli except that lactococci are unable to utilize cytosine.

Cytidine↗
PubMed1993

Octopamine immunoreactive neurons in the fused central nervous system of spiders.

Using antisera directed against octopamine (OA), we identified and mapped octopamine-immunoreactive (OA-ir) neurons and their projections in the fused, central ganglion complex of wandering spiders, Cupiennius salei. Labeled cell bodies are concentrated in the subesophageal ganglion complex (SEG) where they are arranged serially in ventral, midline clusters. OA-ir processes from these cells project dorsally. Some neurites end close to segmental septa; others merge into longitudinal tracts connecting the neuromeres. Labeled collaterals leaving these tracts project into peripheral neuropil. In the brain, OA-ir somata were found only in the two cheliceral hemiganglia, where a cluster of 4-5 relatively large cells (soma diameter 25 microns) lies next to a group of small somata (diameter < 10 microns). Neurites originating from the large somata descend into the SEG and merge into longitudinal tracts. The central body of the brain contains profuse ascending projections. Except for fine varicosities that are confined to the roots of nerves, we found no OA-ir fibers leaving the central nervous system (CNS). Within the CNS, however, OA-ir varicosities are concentrated in neuropil and near hemolymph spaces. This distribution suggests that OA acts as a neurotransmitter and/or local neuromodulator at central synapses, while it is also released into the hemolymph and presumably acts hormonally at peripheral sites. Using high-pressure liquid chromatography measurements, the hemolymph was in fact found to contain 12-40 nM of free octopamine.

Animals↗
PubMed1993

CA 494--a new tumor marker for the diagnosis of pancreatic cancer.

In 59 patients with ductal pancreatic cancer the monoclonal antibody (MAb) BW 494, which detects the CA 494 glycoprotein antigen, was analyzed in comparison with the reference tumor markers CA 19-9 and CEA. Eighty-one patients with non-pancreatic malignancies of the gastrointestinal (GI) tract, 95 with chronic pancreatitis, 124 with benign non-pancreatic GI diseases, 30 with diabetes mellitus (type I or type II) and 114 healthy blood donors served as controls. The sensitivity of pancreatic cancer was 90%, 44% and 90% for CA 19-9, CEA and CA 494, respectively. In chronic pancreatitis, as the most important control population for pancreatic cancer, the specificity was 85%, 72% and 94% for CA 19-9, CEA and CA 494, respectively.

Adolescent↗
PubMed1993

DNA specificity of Escherichia coli deoP1 operator-DeoR repressor recognition.

We have studied the importance of the specific DNA sequence of the deo operator site for DeoR repressor binding by introducing symmetrical, single basepair substitutions at all positions in the deo operator and tested the ability of these variants to titrate DeoR in vivo. Our results show that a 16 bp palindromic sequence constitutes the deo operator. Positions outside this palindrome (positions +/- 9, +/- 10) can be changed without any major effect on DeoR binding. Most of the central 6-8 bp of the palindrome (positions +/- 1, +/- 2, +/- 3) can be substituted with other nucleotides with no or only minor effects on DeoR binding, while changes at position +/- 4 and +/- 5 give a more heterogeneous response. Finally, changes at positions +/- 6, +/- 7 and +/- 8 severely disrupt DeoR binding.

Bacterial Proteins↗
PubMed1993

Functional properties of mechanosensitive units from the chicken ureter in vitro.

A total of 66 mechanosensitive units was recorded from an in vitro preparation of the chicken ureter with attached nerves. They were classified into three groups according to their functional properties. U-1 units (30% of the units) responded to contractions of the ureter and exhibited very low spontaneous activity (mean: 0.1 Hz). They had low average thresholds to distension (5.8 mmHg or 0.77 kPa), responded immediately to mechanical stimuli without exhibiting after discharges to strong stimuli. Their function might be the monitoring of peristalsis. U-2 units (38%) did not respond to contractions and had irregular spontaneous activities (mean: 0.7 Hz). They exhibited a high average activation threshold to distension (42.6 mmHg or 5.73 kPa) and after discharges to strong mechanical stimuli. U-2 units might be involved in nociception. U-G units (32%) were characterized by their long-latency response to any mechanical stimulus used and had a regular high ongoing activity (mean: 2.2 Hz). The pressure thresholds cover the whole range of the U-1 and U-2 units (5 to 70 mmHg or 0.67-9.33 kPa) with an average of 31.5 mmHg or 4.2 kPa. The peak discharge to pressure stimuli occurred much later than in the U-1 and U-2 units and they exhibited pronounced after discharges. Some U-G units were inhibited by mechanical stimuli. U-G units had large receptive fields sometimes covering the entire ureter. The units responded to electrical stimulation of the ureter after a variable latency, indicating that they were synaptically driven. In addition, a late long-lasting response to electrical stimulation was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Minimal Requirements for Exponential Growth of Lactococcus lactis.

A minimal growth medium containing glucose, acetate, vitamins, and eight amino acids allowed for growth of Lactococcus lactis subsp. lactis, with a specific growth rate in batch culture of mu = 0.3 h. With 19 amino acids added, the growth rate increased to mu = 0.7 h and the exponential growth phase proceeded until high cell concentrations were reached. We show that morpholinepropanesulfonic acid (MOPS) is a suitable buffer for L. lactis and may be applied in high concentrations.

Journal Article↗
PubMed1992

Substance P and calcitonin gene-related peptide in the ureter of chicken and guinea-pig: distribution, binding sites and possible functions.

To elucidate the possible functional significance of sensory neuropeptides in visceral organs of mammals and birds the distribution, binding sites and the effects on ureteric peristalsis of substance P and calcitonin gene-related peptide (CGRP) were investigated in the ureter of guinea-pigs and chickens. In the guinea-pig numerous substance P and CGRP-immunoreactive fibres were located in the adventitia, smooth muscle layer, submucosa and occasionally in the epithelium. Varicose peptidergic fibres were often found on blood vessels. Binding sites for substance P were associated with blood vessels and epithelium in the following density order: venules greater than epithelium greater than arterioles. The highest density of CGRP binding sites was detected on the smooth muscle; venules and arterioles expressed moderate binding. The peristalsis frequency of the isolated ureter of the guinea-pig was increased by neurokinin A and substance P, whereas CGRP inhibited ureteric motility. In the chicken the immunoreactivity to substance P and CGRP was less pronounced. Immunoreactive fibres were found in the submucosa close to the epithelium and around ureteric ganglion cells. Correspondingly, substance P binding sites were located in the epithelium and in ureteric ganglia; however, specific CGRP binding was restricted to large blood vessels. In the chicken none of the sensory neuropeptides affected ureteric motility. Only high doses of the sensory neurotoxin capsaicin (greater than 10 microM) repeatedly produced a non-specific inhibitory effect, similar to that found in a capsaicin-desensitized guinea-pig ureter preparation. The data suggest that in the guinea-pig ureter sensory neuropeptides play a modulatory role in the regulation of ureteric motility and might have vascular and epithelial functions. In the chicken, substance P might be involved in the regulation of epithelial function and modulation of ganglionic transmission. The physiological or pathophysiological role of sensory neuropeptides and the efferent functions of afferent fibres appears to be much better developed in the guinea-pig than in the chicken.

Animals↗
PubMed1991

deoP1 promoter and operator mutants in Escherichia coli: isolation and characterization.

Plasmid DNA containing deoP1, one of the two major promoters of the deo operon, has been mutagenized using hydroxylamine, and promoter down-mutations and operator mutations were selected. The isolated mutants are all located within a 16 bp palindromic sequence containing the -10 region of deoP1. The results show that RNA polymerase and DeoR repressor compete for the same DNA target. The deoP1 promotor activity is dependent on a TG motif one base pair upstream of the -10 consensus sequence. The sequence of the deo operator site was further verified by use of a synthetic linker.

Base Sequence↗
PubMed1990

Paralytic shellfish poisoning: a case report and serial electrophysiologic observations.

We report serial electrophysiologic observations in a patient with acute bulbar and respiratory paralysis following ingestion of saxitoxin-contaminated clams. Prolonged distal motor and sensory latencies, slowed conduction velocities, and moderately diminished amplitudes were present at the outset. All values returned to normal over 5 days. These findings, the result of incomplete sodium channel blockade, distinguish paralytic shellfish poisoning from most other acute paralytic illnesses.

Animals↗
PubMed1990

Case management: planning and coordinating strategies.

In summary, planning a case management system involves moving through sequenced stages; namely, (1) a developmental stage; (2) a phase-in stage; and (3) an operational stage. During the developmental stage, before processes have been formalized, attention is given to the political realities, the formation of a representative planning group, formulating goals and objectives, obtaining administrative support, information gathering, assessment of needs, resource procurement, program structure and design, selection of a case management model, participatory decision making, determining organizational fit, and beginning networking. During the phase-in or early implementation state, the formal stage of system introduction, attention is given to the establishment of interorganizational relationships, contracting for services, job descriptions, work assignments, training, problem solving, and conflict resolution. During the operational or full implementation stage, a period when the system should become more stabilized, attention needs to be given to managing movement of the client through the system, the flow of information, program updating, quality assurance, recordkeeping, resource management, evaluating, and system refurbishing. In practice, these stages will interact and overlap. Closure, if the system is to remain viable and open to change, should never occur.

Aged↗
PubMed1989

Single and double loop formation when deoR repressor binds to its natural operator sites.

Distal effects on the in vivo repression of the deo operon are thought to be mediated by the deoR repressor with DNA loop formation. Such loops are easily observed by electron microscopy when the oligomeric deoR repressor is added to a DNA fragment carrying the three genetically defined operators at their chromosomal distances. Upon binding of deoR to any two operators, single loops are formed, 280, 600, and 880 bp in size. With the deo operon, double loops are also formed, which are the combination of the 280 bp and 600 bp loops and the result of simultaneous binding of the protein to its three sites. The formation of both single and double loops is consistent with the long-range effects observed in vivo and with the cooperative involvement of all three operator sites in the repression.

Binding Sites↗
PubMed1989

Purification and characterization of the deoR repressor of Escherichia coli.

The deoR gene, which encodes the deor repressor protein in Escherichia coli, was fused to the strong Ptrc promoter in plasmid pKK233-2. The Ptrc promoter is kept repressed by lacI repressor to prevent cell killing. Induction of the Ptrc--deoR fusion plasmid resulted in the accumulation of 4% of the soluble protein as deoR protein. The deoR repressor protein was purified to 80% purity using conventional techniques; it has a mass of 28.5 kd and appears to exist as an octamer in solution. The deoR repressor is shown by DNase I footprinting to bind to the 16 bp palindromic sequence in the Pribnow box region of the deoP1 promoter. Also, the deoR repressor binds cooperatively in vitro to a DNA template with two deoR binding sites separated by 224 bp in keeping with the conclusion from genetic experiments that more than one operator is required for efficient repression of the deo operon.

Bacterial Proteins↗
PubMed1989

CRP/cAMP- and CytR-regulated promoters in Escherichia coli K12: the cdd promoter.

Transcriptional regulation of the deoP2 promoter by the cyclic AMP/cyclic AMP receptor protein complex (cAMP/CRP) and the CytR repressor requires two high-affinity CRP targets located around -41 and -93 bp preceding the start site for transcription. Here we report the structure of cddP, another CRP/CytR-regulated promoter. In common with what was found in deo, the cdd promoter also contains multiple CRP targets. Thus, using the DNasel footprinting procedure, tandem CRP binding sites were identified around -41 and -93. These findings support a general model for CytR binding and CytR regulation, in which (i) CytR and the CRP/cAMP complex bind to similar or identical targets, (ii) two or more targets are necessary for proper binding of CytR to a promoter region, and (iii) CytR represses transcription by antagonizing cAMP/CRP activation.

Base Sequence↗