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Biomedical subjects

F Buck

Publications and source records attributed to F Buck.

At least 55 records · Page 3Linked to original sources
PubMed1992

Isolation of monocytic serine esterase and evaluation of its proteolytic activity.

Monocytes are characterized by high activity of alpha-naphthyl acetate esterase (ANAE), distinguishing them from all other blood cells. The physiological function of this monocyte marker enzyme has not yet been elucidated. In this study ANAE's potential proteolytic activity was analyzed because serine esterases/proteases can function as effector molecules in cell-mediated cytotoxicity and because monocytes-macrophages are known to exert cytotoxic effects on tumor cells. This enzyme was purified from the monocytic cell line U-937 by preparative isoelectric focusing and a three-step high-performance liquid chromatography that conserved its catalytic activity. It has a molecular mass of 60 kd, and partial amino acid sequence revealed that the enzyme is not identical to known serine esterases/proteases. The purified enzyme failed to digest a couple of peptides, indicating lack of protease activity. In addition, the esterolytic activity of ANAE was not inhibited by protease inhibitors. The isolation and purification of ANAE enable further studies concerning its function in monocytes-macrophages and its relation to monocytic cytotoxicity.

Amino Acid Sequence↗
PubMed1992

Epigenetic activation of Gi-2 protein, the product of a putative protooncogene, mediates tumor promotion in vitro.

Promotion of 'initiated' JB6 epidermal cells to the tumor phenotype can be effected by 12-O-tetradecanoylphorbol-13-acetate treatment, by stimulation of epidermal growth factor (EGF) receptor activity with EGF or transforming growth factor alpha and by exposure to the isoquinoline derivative H7. When these cells were incubated with pertussis toxin (PTX), induction of anchorage-independent growth by all four promoting substances was suppressed. The inhibition is specific since cell proliferation is not affected, suggesting that activation of a Gi protein is essential for promotion of the epidermal cells. This interpretation is strongly supported by the observation that the wasp poison mastoparan, which is known to mimic receptor-mediated activation of certain Gi proteins, also promoted anchorage independence. Immunological data and partial amino acid sequence analysis of ADP-ribosyl alpha i isolated from PTX-treated JB6 cells indicate that a Gi-2 protein is a mediator to tumor promotion in this system. The inhibitory action of 4-bromophenacyl bromide may point to a coupling of the Gi protein to phospholipase A2. From our data we infer that promoters induce the tumor phenotype in 'initiated' JB6 epidermal cells by activating epigenetically the same Gi protein that in a number of adrenal and ovarian tumors appears to be persistently activated by mutational events.

Adrenal Gland Neoplasms↗
PubMed1991

Atrial natriuretic peptide (ANP) is a high-affinity substrate for rat insulin-degrading enzyme.

A cytosolic protein specifically binding to and degrading atrial natriuretic peptide (ANP) was purified from rat brain homogenate. Based on partial amino acid sequences and enzymatic properties, this protein with an apparent molecular mass of 112 kDa has been identified as the rat insulin-degrading enzyme (IDE). In addition to the known substrates, insulin and transforming-growth-factor alpha IDE binds also with high affinity (apparent Kd 60 nM) to ANP. Competition studies with structural variants of ANP demonstrate that both the C terminus and the disulfide loop of the molecule are essential for high-affinity binding. The data suggest that IDE might be involved in the cellular processing and/or metabolic clearance of ANP.

Affinity Labels↗
PubMed1991

Isolation, sequence analysis, and physiological properties of enkephalins in the nervous tissue of the shore crab Carcinus maenas L.

[Leu]- and [Met]enkephalin from thoracic ganglia of the shore crab Carcinus maenas have been purified to homogeneity by a reversed-phase HPLC procedure. Automated gas-phase sequencing revealed a primary structure identical with that of enkephalins from vertebrates. The immunoreactive material in different parts of the nervous system of C. maenas was measured by RIA. Isolated crustacean [Leu]enkephalin shows physiological properties identical to synthetic [Leu]enkephalin in our bioassay. The enkephalinergic effect has been monitored by two bioassays: (i) inhibition of release of crustacean hyperglycemic hormone from isolated eyestalks of C. maenas, and (ii) decreased effect on blood glucose level in the fiddler crab Uca pugilator. The decrease in blood sugar level induced by [Leu]enkephalin was antagonized by naloxone, indicating a receptor-mediated process. In sequence analysis, the thoracic ganglion yielded 3 pmol of [Leu]enkephalin and approximately 1 pmol of [Met]enkephalin. The presence of enkephalins in crustaceans shows an earlier phylogenetic origin of opioids than presumed.

Journal Article↗
PubMed1991

Is the p29 protein involved in the rapid regulation of phosphoenolpyruvate carboxykinase (GTP)?

It has been postulated that a protein with a molecular mass of 29,000 daltons (p29), which copurifies with hepatic phosphoenolpyruvate (P-enolpyruvate) carboxykinase, forms a complex with the enzyme and stabilizes its sensitivity to Mn2+ activation by protecting critical sulfhydryl groups from oxidation (Brinkworth, R. I., Hanson, R. W., Fullin, F. A., and Schramm, V. L. (1981) J. Biol. Chem. 256, 10795-10802). In this paper we demonstrate that p29 is not only expressed in tissues which contain high amounts of P-enolpyruvate carboxykinase, such as liver and kidney, but also in brain and muscle, which have no gluconeogenic function. Furthermore, p29 is expressed in rat liver prenatally, whereas P-enolpyruvate carboxykinase is induced only after birth. The effect of p29 to protect P-enopyruvate carboxykinase against aerobic oxidation during in vitro incubation was also observed for ovalbumin and bovine albumin. Peptide sequencing of the p29 and search in a protein data bank revealed a high homology to the muscle-specific subunit of human phosphoglycerate mutase (EC 2.7.5.3). Determination of the enzyme activity confirms the identification of the p29 as the rat liver isoform of phosphoglycerate mutase. Taking all these findings together, it is concluded that this protein has no specific effect on P-enolpyruvate carboxykinase.

Amino Acid Sequence↗
PubMed1991

Characterization of N- and C-terminal deletion mutants of the rat serotonin HT2 receptor in Xenopus laevis oocytes.

Deletion mutants of the serotonin HT2 receptor have been constructed in which the N- and C-terminal sequences have been gradually removed. The mutant constructs were assayed for their biological activity by electrophysiological measurements in Xenopus oocytes previously injected with the respective in vitro synthesized cRNAs. No significant loss of biological activity was observed when either the extracellular N-terminal sequence, including all potential glycosylation sites, up to the beginning of the first transmembrane domain or the C-terminal sequence up to cystein residue 397, which is conserved in most G-protein coupled receptor known so far, was deleted from the serotonin HT2 receptor constructs.

Amino Acid Sequence↗
PubMed1991

cDNA cloning and characterization of human monocyte/macrophage serine esterase-1.

Human monocyte/macrophage serine esterase (HMSE), commonly known as acid esterase or alpha-naphthylacetate esterase, comprises a group of five enzyme variants that can be distinguished by their isoelectric points from esterase variants of the other normal human blood cell populations. A cDNA for one of the monocytic enzyme variants (HMSE1) was cloned from a U-937 lambda gt11 cDNA library by screening with an oligonucleotide mixture designed according to amino acid sequence data of the purified enzyme. The cDNA contains 1,727 bp with an open reading frame of 1,512 bp coding for a protein of 503 amino acid residues. HMSE1 cDNA represents the first cloned monocyte/macrophage-specific serine esterase and its sequence shows up to 77% homology to other known serine esterases of different species. The amino acid composition of the putative active site of HMSE1 as deduced from the nucleotide sequence corresponds with the active sites of other serine esterases but not with the active sites of serine proteases. Hybridization of the cDNA with RNA of separated normal blood cell populations and hematopoietic cell lines shows restricted expression within the monocyte/macrophage lineage.

Amino Acid Sequence↗
PubMed1991

Purification and cDNA sequencing of an oleate-selective acyl-ACP:sn-glycerol-3-phosphate acyltransferase from pea chloroplasts.

The soluble acyl-ACP:sn-glycerol-3-phosphate acyltransferase from chloroplasts of chilling-sensitive and -resistant plants differ in their fatty acid selectivity. Enzymes from resistant plants discriminate against non-fluid palmitic acid and select oleic acid whereas the acyltransferase from sensitive plants accepts both fatty acids. To use this difference for improving plant chilling resistance by biotechnology the gene for an oleate-selective enzyme is required. Therefore, the oleate-selective enzyme from pea seedlings was purified to apparent homogeneity. Tryptic peptides of internal origin were sequenced. Polyclonal antibodies raised in rabbits were used for an immunological screening of a pea leaf cDNA expression library in lambda gt11. A positive clone of 1800 bp was selected showing an open reading frame which codes for 457 amino acids. The deduced amino acid sequence coincides perfectly with the tryptic sequences. A tentative assignment of the processing site was made which divides the preprotein into a mature protein of 41 kDa in accordance with experimental findings and a transit peptide of 88 amino acids. At present the comparison between a selective (pea) and an unselective (squash) acyltransferase sequence does not provide a clue for recognizing the structural differences resulting in different selectivities.

1-Acylglycerophosphocholine O-Acyltransferase↗
PubMed1991

Neuropeptide hormone receptors: strategies for identification.

Study of the structure--function relationship of neuropeptide hormone receptors presents a number of technical difficulties associated with the isolation of a given receptor protein in a purified form. A variety of molecular approaches has enabled corresponding cDNA clones to be isolated without the need to embark on protein purification procedures. However, the molecular cloning approach requires that appropriate tools for identifying cDNAs encoding the respective receptor be available. Strategies designed to address this problem will be discussed and include functional expression of neuropeptide hormone receptors in frog oocytes, hybrid depletion and inactivation of receptor-encoding mRNAs by RNase H digestion, polymerase chain reaction (PCR) amplification of cDNAs encoding putative receptors, and expression of transfected receptor genes in cell cultures followed by identification using a cell sorter.

Amino Acid Sequence↗
PubMed1990

[Weakness, bone pain].

A 57-year-old man complains of a reduced exercise tolerance and bone pain for years. There is a past medical history of hyperuricemia and anemia of unknown etiology. In a routine examination a leukocytosis of 64,000 per ml, increased number of thrombocytes and an elevated level of lactic dehydrogenase is found. The suspected diagnosis of myeloproliferative disorder, in the present case of chronic myelogenous leukemia, is confirmed by a diminished neutrophil alkaline phosphatase activity, bone marrow biopsy and the demonstration of the Ph1 (Philadelphia) chromosome. Magnetic resonance imaging reveals an infiltration of the bone marrow. After the initial treatment with busulfan the white cell count is lowered.

Bone Diseases↗
PubMed1989

NMR study of the structural changes induced in the E. coli lac promoter by the specific binding of the CAP protein.

We have studied the binding of the CAP protein to an 18 base pair lac promoter sequence comprising the core of the CAP recognition sequence. Specific binding of this sequence was established by competition binding assays and comparison of the relative affinities of a number of lac promoter, lac operator, and unspecific sequences of different lengths. The effect of the binding of CAP to the 18 base pair promoter sequence and, for comparison, to an 18 base pair symmetric operator and an oligonucleotide of unrelated sequence have been studied by 1H NMR. Binding of CAP does not bring about any changes in the chemical shift values of the imino proton resonances of the DNA, but causes the selective line broadening of two of the resonances. The comparison of these data with results of gel retardation assays published previously (1) allows the identification and localization of a kink induced in the DNA by the CAP binding to its specific site on the lac promoter.

Bacterial Proteins↗
PubMed1989

H NMR study of a complex between the lac repressor headpiece and a 22 base pair symmetric lac operator.

A complex between the lac repressor headpiece and a fully symmetric tight-binding 22 bp lac operator was studied by 2D NMR. Several 2D NOE spectra were recorded for the complex in both H2O and 2H2O. Many NOE cross-peaks between the headpiece and DNA could be identified, and changes in the chemical shift of the DNA protons upon complex formation were analyzed. Comparison of these data with those obtained for a complex between the headpiece and a 14 bp half-operator, studied previously [Boelens, R., Scheek, R. M., Lamerichs, R. M. J. N., de Vlieg, J., van Boom, J. H., & Kaptein, R. (1987) in DNA-ligand interactions (Guschlbauer, W., & Saenger, W., Eds.) pp 191-215, Plenum, New York], shows that two headpieces form a specific complex with the 22 bp lac operator in which each headpiece binds in the same way as found for the 14 bp complex. The orientation of the recognition helix in the major groove of DNA in these complexes is opposite with respect to the dyad axis to that found for other repressors.

Base Composition↗
PubMed1989

A single-step purification procedure and partial amino acid sequence analysis of picomole amounts of the rat T cell alloantigen RT6.2.

A single-step immunoaffinity purification procedure for the rat T cell marker RT6.2 is described which permits the isolation of microgram quantities of protein from the RT6.2+ T-T hybridoma EpD3. The amino terminus was sequenced directly from a polyvinylidene (PVDF) membrane blot prepared after SDS-PAGE. Further internal sequence data were obtained from peptides generated from purified RT6.2 digested with different endoproteases and separated by reverse-phase micro-HPLC. A computer search in data banks did not reveal any significant homology to other proteins.

ADP Ribose Transferases↗
PubMed1987

One- and two-dimensional 1H-NMR investigations of two 19-base-pair analogues of the tet operator.

Two 19-base-pair oligodeoxynucleotides, analogues of one of the operators which specifically bind the repressor protein in the regulatory part of the transposon Tn10 tetracycline-resistance (tet) determinant, have been studied by 1H-NMR spectroscopy. The analogues contain a mismatch in the central base pair of the double helix (T.T or A.A). The imino protons have been assigned to the base pairs by one-dimensional NOE measurements, and the thermally induced transition from the duplex to the single strand has been followed. The cytidine amino resonances have been assigned by means of two-dimensional NOE spectroscopy in H2O. Two-dimensional phase-sensitive NOE and magnitude-correlated spectra have been recorded in 2H2O; all nonexchangeable protons, with the exception of some of H5', H5" protons, have been assigned. The NMR data made it possible to carry out a qualitative analysis of the structures of both oligodeoxynucleotides. The general structures close to B-DNA, show irregularities in the mismatch areas.

Base Sequence↗
PubMed1987

15N labeling of oligodeoxynucleotides for NMR studies of DNA-ligand interactions.

The amino protons of 15N-labeled DNA were studied as a possible structural probe in NMR investigations of the interaction of DNA with various ligands. Since the imino protons are located in the center of the double helix, and variations of their chemical shift values are difficult to interpret in terms of structural changes, these probes are not very useful. Instead, amino protons are located in the major or minor groove of the DNA and are often directly involved in the binding of a ligand. For a selective probing 4-15NH2-2'-deoxycytidine and 6-15NH2-2'-deoxyadenosine were obtained by chemical synthesis. The labeled nucleosides were introduced in distinct positions of oligodeoxynucleotides by large-scale DNA synthesis. Direct 15N NMR and 1H-15N multiple quantum NMR were applied to detect the corresponding 15N labels or protons attached to the 15N labels. Chemical shift values for the cytidine and the adenosine amino nitrogen and proton resonances of a symmetric 18 base pair lac operator sequence are reported.

Adenine↗
PubMed1987

Sarcoidosis and amyloidosis. Is the association causal or co-incidental?

During 1950-83, 52,371 autopsies were done at the Los Angeles County Hospital; 503 cases showing amyloidosis, seven of these also had sarcoidosis, and three also had rheumatoid arthritis. These associations could be coincidental or it is just possible that there is a causal aetiological relationship. It is hoped that other workers will review their material, and, in particular, to assess whether there is an increased incidence of amyloidosis in sarcoidosis.

Adult↗
PubMed1986

Base substitution mutants of the lac operator: in vivo and in vitro affinities for lac repressor.

16 single-site mutations and a 1-bp deletion in the lac operator have been cloned and examined with regard to repressor binding. A 13-bp, central 'core' operator sequence, bp 5-17 of the natural operator, was also synthesized and cloned. Repressor affinity was assessed in vivo by quantitating the level of beta-galactosidase activity resulting from chromosomal operon derepression and in vitro by measuring the stability of repressor-operator complexes. Our results support the general conclusion that the repressor-operator interaction is asymmetric, particularly across the center of the operator sequence, with little or no specific contact at position 12. Some sequence changes in the right side of the operator markedly reduced repressor affinity, indicating that although binding to this half of the sequence has been suggested to be less important than the left half, it still significantly contributes to the binding affinity.

Cloning, Molecular↗