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B Kempf

Publications and source records attributed to B Kempf.

18 recordsLinked to original sources
PubMed1996

A novel amidohydrolase gene from Bacillus subtilis cloning: DNA-sequence analysis and map position of amhX.

The nucleotide sequence of a new Bacillus subtilis gene (amhX) was determined that encodes a protein (AmhX) with strong sequence identity to amidohydrolases from both plant and bacterial species and a carboxypeptidase from the archaeon Sulfolobus sulfataricus. The amhX gene encodes a hydrophilic polypeptide of 383 amino acids with a molecular mass of 41.5 kDa. The amhX gene was overexpressed in E. coli by using the T7 RNA polymerase/promoter system and the transcription initiation sites for the amhX mRNAs in B. subtilis were determined by primer extension analysis. Chromosomal amhX mutations were constructed by marker replacement and the amhX gene was positioned at 25 degrees on the genetic and physical map of the B. subtilis chromosome.

Amidohydrolases↗
PubMed1996

Three transport systems for the osmoprotectant glycine betaine operate in Bacillus subtilis: characterization of OpuD.

The accumulation of the osmoprotectant glycine betaine from exogenous sources provides a high degree of osmotic tolerance to Bacillus subtilis. We have identified, through functional complementation of an Escherichia coli mutant defective in glycine betaine uptake, a new glycine betaine transport system from B. subtilis. The DNA sequence of a 2,310-bp segment of the cloned region revealed a single gene (opuD) whose product (OpuD) was essential for glycine betaine uptake and osmoprotection in E. coli. The opuD gene encodes a hydrophobic 56.13-kDa protein (512 amino acid residues). OpuD shows a significant degree of sequence identity to the choline transporter BetT and the carnitine transporter CaiT from E. coli and a BetT-like protein from Haemophilus influenzae. These membrane proteins form a family of transporters involved in the uptake of trimethylammonium compounds. The OpuD-mediated glycine betaine transport activity in B. subtilis is controlled by the environmental osmolarity. High osmolarity stimulates de novo synthesis of OpuD and activates preexisting OpuD proteins to achieve maximal glycine betaine uptake activity. An opuD mutant was constructed by marker replacement, and the OpuD-mediated glycine betaine uptake activity was compared with that of the previously identified multicomponent OpuA and OpuC (ProU) glycine betaine uptake systems. In addition, a set of mutants was constructed, each of which synthesized only one of the three glycine betaine uptake systems. These mutants were used to determine the kinetic parameters for glycine betaine transport through OpuA, OpuC, and OpuD. Each of these uptake systems shows high substrate affinity, with Km values in the low micromolar range, which should allow B. subtilis to efficiently acquire the osmoprotectant from the environment. The systems differed in their contribution to the overall glycine betaine accumulation and osmoprotection. A triple opuA, opuC, and opuD mutant strain was isolated, and it showed no glycine betaine uptake activity, demonstrating that three transport systems for this osmoprotectant operate in B. subtilis.

Amino Acid Sequence↗
PubMed1996

Synthesis of the osmoprotectant glycine betaine in Bacillus subtilis: characterization of the gbsAB genes.

Synthesis of the osmoprotectant glycine betaine from the exogenously provided precursor choline or glycine betaine aldehyde confers considerable osmotic stress tolerance to Bacillus subtilis in high-osmolarity media. Using an Escherichia coli mutant (betBA) defective in the glycine betaine synthesis enzymes, we cloned by functional complementation the genes that are required for the synthesis of the osmoprotectant glycine betaine in B. subtilis. The DNA sequence of a 4.1-kb segment from the cloned chromosomal B. subtilis DNA was established, and two genes (gbsA and gbsB) whose products were essential for glycine betaine biosynthesis and osmoprotection were identified. The gbsA and gbsB genes are transcribed in the same direction, are separated by a short intergenic region, and are likely to form an operon. The deduced gbsA gene product exhibits strong sequence identity with members of a superfamily of specialized and nonspecialized aldehyde dehydrogenases. This superfamily comprises glycine betaine aldehyde dehydrogenases from bacteria and plants with known involvement in the cellular adaptation to high-osmolarity stress and drought. The deduced gbsB gene product shows significant similarity to the family of type III alcohol dehydrogenases. B. subtilis mutants with defects in the chromosomal gbsAB genes were constructed by marker replacement, and the growth properties of these mutant strains in high-osmolarity medium were analyzed. Deletion of the gbsAB genes destroyed the choline-glycine betaine synthesis pathway and abolished the ability of B. subtilis to deal effectively with high-osmolarity stress in choline- or glycine betaine aldehyde-containing medium. Uptake of radiolabelled choline was unaltered in the gbsAB mutant strain. The continued intracellular accumulation of choline or glycine betaine aldehyde in a strain lacking the glycine betaine-biosynthetic enzymes strongly interfered with the growth of B. subtilis, even in medium of moderate osmolarity. A single transcription initiation site for gbsAB was detected by high-resolution primer extension analysis. gbsAB transcription was initiated from a promoter with close homology to sigma A-dependent promoters and was stimulated by the presence of choline in the growth medium.

Alcohol Dehydrogenase↗
PubMed1995

OpuA, an osmotically regulated binding protein-dependent transport system for the osmoprotectant glycine betaine in Bacillus subtilis.

Exogenously provided glycine betaine can efficiently protect Bacillus subtilis from the detrimental effects of high osmolarity environments. Through functional complementation of an Escherichia coli mutant deficient in glycine betaine uptake with a gene library from B. subtilis, we have identified a multicomponent glycine betaine transport system, OpuA. Uptake of radiolabeled glycine betaine in B. subtilis was found to be osmotically stimulated and was strongly decreased in a mutant strain lacking the OpuA transport system. DNA sequence analysis revealed that the components of the OpuA system are encoded by anoperon (opuA) comprising three structural genes: opuAA, opuAB, and opuAC. The products of these genes exhibit features characteristic for binding protein-dependent transport systems and in particular show homology to the glycine betaine uptake system ProU from E. coli. Expression of the opuA operon is under osmotic control. The transcriptional initiation sites of opuA were mapped by high resolution primer extension analysis, and two opuA mRNAs were detected that differed by 38 base pairs at their 5' ends. Synthesis of the shorter transcript was strongly increased in cells grown at high osmolarity, whereas the amount of the longer transcript did not vary in response to medium osmolarity. Physical and genetic mapping experiments allowed the positioning the opuA operon at 25 degrees on the genetic map of B. subtilis.

Amino Acid Sequence↗
PubMed1995

The osmoprotectant proline betaine is a major substrate for the binding-protein-dependent transport system ProU of Escherichia coli K-12.

The ProP and ProU transport systems of Escherichia coli mediate the uptake of several osmoprotectants including glycine betaine. Here we report that both ProP and ProU are involved in the transport of the potent osmoprotectant proline betaine. A set of isogenic E. coli strains carrying deletions in either the proP or proU loci was constructed. The growth properties of these mutants in high osmolarity minimal media containing 1 mM proline betaine demonstrated that the osmoprotective effect of this compound was dependent on either an intact ProP or ProU uptake system. Proline betaine competes with glycine betaine for binding to the proU-encoded periplasmic substrate binding protein (ProX) and we estimate a KD of 5.2 microM for proline betaine binding. This value is similar to the binding constant of the ProX protein determined previously for the binding of glycine betaine (KD of 1.4 microM). Our results thus demonstrate that the binding-protein-dependent ProU transport system of E. coli mediates the efficient uptake of the osmoprotectants glycine betaine and proline betaine.

Amino Acid Transport Systems↗
PubMed1995

In vitro investigation of various attachments for overdentures on osseointegrated implants.

Primary splinting of fixtures with bar attachments has proved to be clinically effective for overdentures on osseointegrated implants in the edentulous mandible. However, in vitro investigations indicate that a more favourable force transfer to the individual fixtures can be attained by secondary splinting of four implants with conical double crowns. This observation could be confirmed in the present study by measurement of implant deflection in a model of the mandible by means of interferometric optical holography. Telescope crowns with conical interface brought about functionally more even movement characteristics of the implants compared to rigid or jointed bar attachments. Moreover, the conical double crowns always ensured a stable position of the denture in contrast to jointed attachments. These results underscore the need to test conical double crowns clinically as anchors for overdentures on osseointegrated implants.

Crowns↗
PubMed1994

Interactions of the nucleoid-associated DNA-binding protein H-NS with the regulatory region of the osmotically controlled proU operon of Escherichia coli.

The Escherichia coli hns gene encodes the abundant nucleoid-associated DNA-binding protein H-NS. Mutations in hns alter the expression of many genes with unrelated functions and result in a derepression of the proU operon (proVWX) without abolishing the osmotic control of its transcription. We have investigated the interactions of H-NS with the proU regulatory region by deletion analysis of cis-acting sequences, competitive gel retardation assays, and DNase I footprinting. The negative effect of H-NS on proU transcription was mediated by cis-acting sequences within proV but did not depend on the presence of a curved DNA segment upstream of the proU-35 region previously characterized as a target for H-NS binding in vitro. We detected a 46-base pair high affinity H-NS binding region downstream of the proU promoter at the 5' end of the proV gene and a complex array of additional H-NS binding sites which suggest the presence of an extended H-NS nucleoprotein complex. Most of the H-NS binding sites were highly A+T-rich and carried stretches of 5 or more consecutive A-T base pairs. The implications of our results for the osmotic regulation of proU transcription are discussed.

Amino Acid Transport Systems↗
PubMed1994

Osmoregulation in Bacillus subtilis: synthesis of the osmoprotectant glycine betaine from exogenously provided choline.

Exogenously provided glycine betaine functions as an efficient osmoprotectant for Bacillus subtilis in high-osmolarity environments. This gram-positive soil organism is not able to increase the intracellular level of glycine betaine through de novo synthesis in defined medium (A. M. Whatmore, J. A. Chudek, and R. H. Reed, J. Gen. Microbiol. 136:2527-2535, 1990). We found, however, that B. subtilis can synthesize glycine betaine when its biosynthetic precursor, choline, is present in the growth medium. Uptake studies with radiolabelled [methyl-14C]choline demonstrated that choline transport is osmotically controlled and is mediated by a high-affinity uptake system. Choline transport of cells grown in low- and high-osmolarity media showed Michaelis-Menten kinetics with Km values of 3 and 5 microM and maximum rates of transport (Vmax) of 10 and 36 nmol min-1 mg of protein-1, respectively. The choline transporter exhibited considerable substrate specificity, and the results of competition experiments suggest that the fully methylated quaternary ammonium group is a key feature for substrate recognition. Thin-layer chromatography revealed that the radioactivity from exogenously provided [methyl-14C]choline accumulated intracellularly as [methyl-14C]glycine betaine, demonstrating that B. subtilis possesses enzymes for the oxidative conversion of choline into glycine betaine. Exogenously provided choline significantly increased the growth rate of B. subtilis in high-osmolarity media and permitted its proliferation under conditions that are otherwise strongly inhibitory for its growth. Choline and glycine betaine were not used as sole sources of carbon or nitrogen, consistent with their functional role in the process of adaptation of B. subtilis to high-osmolarity stress.

Bacillus subtilis↗
PubMed1989

[Creatine kinase risk groups in acute myocardial infarct].

The individual valuation of risks in patients with acute myocardial infarction on the basis of a monitoring of the creatine kinase (CK) is made evident as relevant to practice for the basic medical care. Thereby a classification of risk groups on the basis of CKmax (less than or equal to 23; greater than 23 less than or equal to 40; greater than 40 less than or equal to 60; greater than 60 mumol/l.s) is controlled. The measurement of the ejection fraction global was performed also according to risk groups (greater than 60; greater than 45 less than or equal to 60; greater than 30 less than or equal to 45; less than or equal to 30%). Parallel to this a classification of the patients according to the electrocardiogram (non-Q-wave, Q-wave) was retrospectively performed. The anamnestic information Re-AMI was individually taken into consideration for the evaluation of CKmax. In 2.5% of the patients in comparison to the ejection fraction the risk group classification of CKmax was globally carried out into a higher group without an an principal incorrect evaluation of the risk (slight, middle, high) in the first AMI. No patient with middle or high risk in the first AMI was incorrectly grouped according to the risk group CKmax or ejection fraction global.

Cardiac Output↗
PubMed1984

[Multiple Cancers].

According to the criteria of Warren and Gates the course of disease of 11972 patients--8099 of them carcinomas under treatment in the Universitäts-Strahlenklinik--were examined in respect to multiple malignomas. In 232 cases such multiple malignomas were diagnosed, corresponding to 2.86%. We found more women (3.49%) than men (2.13%). In 165 cases (67.2%) multiple malignomas appeared metachronically, i.e. after an interval of more than 2 years, 76 (32.8%) of those patients were synchronic multiple malignoma--carriers. In women we found with 3.49% significantly more often metachronic manifestations than in men (2.13%). According to synchronic multiple malignomas we stated that they are to be found in males and females in an equal amount. On the average the second malignoma appeared in males and females 10 years after diagnosis of the primary tumor. Localizations of the multiple malignomas as well as combinations of certain tumors are shown and discussed.

Adult↗
PubMed1982

Opioid peptides and blood pressure regulation.

Immunohistochemical and functional studies suggest that opioid peptides are involved in blood pressure regulation. Endorphin and enkephalins contained in the brain seem to exert their effects by mediation of cardiovascular centers in the brain stem. Additional effects on blood pressure may be exhibited by circulating opioid peptides released from the pituitary and adrenal gland which may act at both central and peripheral sites. It is expected that further studies will complete our knowledge about the interaction of opioid peptides and blood pressure regulation and will result in therapeutic applications.

Animals↗