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Biomedical subjects

U Pieper

Publications and source records attributed to U Pieper.

29 records · Page 2Linked to original sources
PubMed1992

Molecular basis for biosynthesis and accumulation of polyhydroxyalkanoic acids in bacteria.

The current knowledge on the structure and on the organization of polyhydroxyalkanoic acid (PHA)-biosynthetic genes from a wide range of different bacteria, which rely on different pathways for biosynthesis of this storage polyesters, is provided. Molecular data will be shown for genes of Alcaligenes eutrophus, purple non-sulfur bacteria, such as Rhodospirillum rubrum, purple sulfur bacteria, such as Chromatium vinosum, pseudomonads belonging to rRNA homology group I, such as Pseudomonas aeruginosa, Methylobacterium extorquens, and for the Gram-positive bacterium Rhodococcus ruber. Three different types of PHA synthases can be distinguished with respect to their substrate specificity and structure. Strategies for the cloning of PHA synthase structural genes will be outlined which are based on the knowledge of conserved regions of PHA synthase structural genes and of the PHA-biosynthetic routes in bacteria as well as on the heterologous expression of these genes and on the availability of mutants impaired in the accumulation of PHA. In addition, a terminology for the designation of PHAs and of proteins and genes relevant for the metabolism of PHA is suggested.

Acyltransferases↗
PubMed1991

Site-directed mutagenesis studies with EcoRV restriction endonuclease to identify regions involved in recognition and catalysis.

Guided by the X-ray structure analysis of a crystalline EcoRV-d(GGGATATCCC) complex (Winkler, in preparation), we have begun to identify functionally important amino acid residues of EcoRV. We show here that Asn70, Asp74, Ser183, Asn185, Thr186, and Asn188 are most likely involved in the binding and/or cleavage of the DNA, because their conservative substitution leads to mutants of no or strongly reduced activity. In addition, C-terminal amino acid residues of EcoRV seem to be important for its activity, since their deletion inactivates the enzyme. Following the identification of three functionally important regions, we have inspected the sequences of other restriction and modification enzymes for homologous regions. It was found that two restriction enzymes that recognize similar sequences as EcoRV (DpnII and HincII), as well as two modification enzymes (M.DpnII and, in a less apparent form, M.EcoRV), have the sequence motif -SerGlyXXXAsnIleXSer- in common, which in EcoRV contains the essential Ser183 and Asn188 residues. Furthermore, the C-terminal region, shown to be essential for EcoRV, is highly homologous to a similar region in the restriction endonuclease SmaI. On the basis of these findings we propose that these restriction enzymes and to a certain extent also some of their corresponding modification enzymes interact with DNA in a similar manner.

Amino Acid Sequence↗
PubMed1990

Genetic engineering, isolation and characterization of a truncated Escherichia coli elongation factor Tu comprising domains 2 and 3.

A deletion mutant of a plasmid born Escherichia coli tufA gene, which codes for a truncated elongation factor Tu comprising domains 2 and 3, has been constructed by genetic engineering. This gene was overexpressed in E. coli, and a polypeptide representing the truncated elongation factor Tu was isolated, purified to near homogeneity, crystallized and characterized physico-chemically as well as biochemically. Circular dichroism spectroscopy and limited tryptic digestion demonstrate that the isolated domain pair 2 and 3 behaves like an independent folding unit which adopts a similar secondary and most likely, tertiary, structure to that present in the intact elongation factor Tu. However, the isolated domain pair 2 and 3 does not interact with aminoacyl-tRNA or the antibiotic kirromycin, two ligands which were shown previously by cross-linking experiments to be in contact with amino acid residues located in domains 1 and 2, and domain 3, respectively. The results suggest that the isolated domain pair 2 and 3 by itself forms too few contacts with these ligands to form a stable complex. Furthermore, the data suggest that domain 1 in intact EF-Tu, in a subtle but nevertheless decisive manner, alters the conformation of the other two domains in such a way that all three domains cooperatively create a high affinity binding site for aminoacyl-tRNA and the antibiotic kirromycin.

Circular Dichroism↗
PubMed1988

Spectroscopic and hydrodynamic studies reveal structural differences in normal and transforming H-ras gene products.

We have recorded the circular dichroism spectra of the cellular and the viral H-ras gene products both in the absence and in the presence of guanine nucleotides and analyzed these spectra in terms of the secondary structure composition of these proteins. It is shown that the GTP complex of the ras proteins has a different secondary structure composition than the GDP complex and, furthermore, that there are differences in the secondary structure of the viral ras protein and the cellular ras protein. We have also recorded and analyzed the circular dichroism spectrum of the isolated guanine nucleotide binding domain of the Escherichia coli elongation factor Tu (EF-Tu), which has been considered as a model for the tertiary structure of the ras proteins [McCormick, F., Clark, B. F. C., LaCour, T. F. M., Kjeldgaard, M., Norskov-Lauritsen, L., & Nyborg, J. (1985) Science (Washington, D.C.) 230, 78-82]. Our data show that the guanine nucleotide binding domain of EF-Tu (30% alpha-helix and 16% beta-pleated sheet for the GDP complex) has quite a different secondary structure composition than the ras proteins (e.g., the cellular ras protein has 47% alpha-helix and 22% beta-pleated sheet for the GDP complex), indicating that the protein core comprising the guanine nucleotide binding site might be similar but that major structural differences must exist at the portion outside this core. Normal and transforming ras proteins also differ slightly in their hydrodynamic properties as shown by sedimentation velocity runs in the analytical ultracentrifuge.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Transformation, Neoplastic↗
PubMed1988

Collagenase activity in cervical tissue of the non-pregnant and pregnant human cervix.

Collagen degradation has been suggested to play an important role in the process of cervical ripening at term. After giving informed consent tissue samples were taken from the posterior lip of 10 non-pregnant women of child-bearing age, 10 women having a termination of pregnancy at 9-12 weeks' gestation and 12 women who had had normal spontaneous deliveries at term. Collagenase activity was measured by a specific technique using native, triple helical collagen. Proteolytic activity was determined with commercially available 14C-methaemoglobin as substrate. For identification of collagen fragments SDS-polyacrylamid-gel-electrophoresis was performed using a modified Laemmli-technique. Collagenase and protease activity were found in all extracts of the different groups. There was a marked increase in enzymatic activities in the postpartum tissue samples suggesting that significant collagen breakdown had taken place. The absence of typical collagen fragments in the SDS-electrophoresis seemed to be due to a very rapid further degradation of the fragments by proteases into small peptides which then readily left the cervix. The present findings confirm the essential role of proteolytic enzymes in cervical dilatation at term.

Adult↗
PubMed1987

Collagen degradation in the pregnant human cervix at term and after prostaglandin-induced cervical ripening.

The role of enzymatic collagen degradation in prostaglandin-induced and physiological cervical ripening was studied by determining collagenase and other proteolytic activity in extracts of cervical biopsies. Collagenase activity was assayed in a highly specific and sensitive system using native collagen type I as substrate. The intracervical application of sulprostone gel prior to termination of 1st trimester pregnancy led to a marked improvement in cervical dilatability. Collagenase and proteolytic activity were found to be significantly higher in postpartum samples than in specimens obtained from the nonpregnant or early pregnant cervix. After sulprostone treatment enzymatic activities were only marginally elevated. Analysis of extractable peptides showed that sub partu collagen was degraded in preference to noncollagenous proteins into very small fragments, whereas in the process of prostaglandin-induced cervical ripening collagen degradation appears to be of minor importance.

Cervix Uteri↗
PubMed1987

The role of collagenases and proteases in prostaglandin-induced cervical ripening.

Tissue samples were taken from the posterior lip of the cervix of 10 non-pregnant women, 10 women having a termination of pregnancy at 9-12 weeks' gestation and 16 women having a termination who had had an intracervical application of either 50 micrograms sulprostone gel or 2 ml 5% tylose 8-12 hours previously. The efficacy of cervical priming was demonstrated objectively by tonometric measurements. Collagenase activity was determined by a new highly specific technique using native, triple helical collagen. Protease activity was measured by a modified Anson-test. For identification of collagen fragments SDS-polyacrylamide electrophoresis was done on the acetic acid soluble fractions. The sulprostone gel induced effective cervical ripening in all of the patients. Collagenase and protease activity were found in all extracts from the different groups, however, PG-pretreatment of the cervix led to no significant increase in enzymatic activities. In addition, the absence of typical collagen cleavage products in the SDS-electrophoresis suggested that no significant collagen breakdown had occurred. In contrast to previously published literature we conclude that enzymatic collagen degradation does not play a predominant role in PG-induced cervical ripening.

Abortifacient Agents, Nonsteroidal↗
PubMed1986

Effect of misidentification on the estimation of breeding value and heritability in cattle.

For daughter groups of 15 test bulls, controls of paternity were performed by using blood group factors and biochemical polymorphisms. Data of incorrectly assigned daughters influenced the estimation of breeding values, heritabilities and correlations for milk performance traits. Formulae are given that show the effects of variable misidentification rates on estimation of breeding values, selection intensities, heritabilities, and genetic gains. For example, for milk fat yield, the genetic gains drop at a misidentification rate of 15% between 8.7% (for h2 = .5) and 16.9% (for h2 = .2) below values attained without misidentifications. Consequently, decreasing misidentification rates in progeny of test bulls can be used to diminish the progeny size per test bull for constant genetic gain, to achieve more precise ranking of all or distinct test bulls according to their "true" breeding values and(or) to increase the number of test bulls by using the same amount of test inseminations and the same precision of ranking. Actions to reduce misidentification rates in cattle populations are discussed.

Animal Identification Systems↗
PubMed1978

[Racial characteristics from the industrial anthropology aspect].

Often industrial-anthropological research cannot be limited to a particular population. Whenever export and/or migration brings people and conditions together which go beyond the usual area of a population characterized by a particular morphological trait, the question arises as to how technical conditions can best be adapted to humans. The automobile seat is used to illustrate to what extent race-specific size and body proportion differences effect technical practice. Since the safety of an automobile is in part dependent upon an efficient adaptation of the vehicle to the user, these investigations have considerable practical importance.

Adult↗
PubMedDate not supplied

How many EF-Tu molecules participate in aminoacyl-tRNA binding?

The stoichiometry of the EF-Tu-GTP-aminoacyl-tRNA complex has been re-determined by a variety of methods, viz gel filtrations, fluorescence titrations, as well as hydrolysis and RNase protection experiments. The results of these experiments clearly demonstrate that one aminoacyl-tRNA interacts with only one EF-Tu-GTP molecule, in agreement with the established view and in contrast to the recently published results by Ehrenberg et al [6].

Binding Sites↗