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Biomedical subjects

M W Chen

Publications and source records attributed to M W Chen.

32 records · Page 2Linked to original sources
PubMed1995

Peptide growth factors in normal and hypertrophied bladder.

The bladder is a dynamic organ that responds to the stress of outlet obstruction by rapidly increasing in mass and cellular content to compensate for increased urethral resistance. If the outlet obstruction is released prior to decompensation, the hypertrophied bladder will shrink, returning to normal size and cellularity. However, with chronic obstruction the bladder will continue to increase in mass, developing drastic alterations in the amount and composition of the extracellular matrix and, ultimately, losing the ability to function. The extensive tissue remodeling associated with each of these changes requires different bladder components to undergo cellular proliferation, cellular hypertrophy, and even programmed cell death (apoptosis). Each of these cellular processes is known to be regulated by various peptides that are referred to as growth factors. Herein we provide an overview of the growth factors that are known to influence the bladder in addition to a variety of experimental animal studies that putatively identify a role of four particular growth factors [basic fibroblast growth factor (bFGF), transforming growth factor-beta (TGF-beta), nerve growth factor (NGF), and epidermal growth factor (EGF)] in the obstructive bladder response.

Animals↗
PubMed1994

Changes in lumbar disc angles of Chinese subjects from upright to flexion.

STUDY DESIGN: This study analyzed the sizes and angles of vertebrae and discs in Chinese subjects. Lateral lumbar spine radiograph films were taken of subjects in upright and 60 degrees flexion positions to evaluate disc height and angle changes. OBJECTIVES: This study sought to gather data regarding the contribution of each segment to lumbar lordotic curvature in upright posture, and angle changes of individual segments from upright to flexion positions. SUMMARY OF BACKGROUND DATA: Although the dimensions of vertebrae and discs have been measured, little has been done regarding in vivo measurements of lumbar lordotic curvature formation, especially during flexion. In addition, no data are available for the Chinese population. METHODS: Five men and six women were studied. The lateral lumbar spine radiograph films were taken for upright and 60 degrees flexion positions. The dimensions of vertebral bodies and discs and lumbosacral angle were measured. RESULTS: The disc heights ranged from 0.7 to 1.2 cm in upright posture. Mean disc angle ranged from 9 degrees to 14 degrees in men and 6 degrees to 13 degrees in women. The lowest three segments (L3-L5) contributed 80% to 90% of the lumbosacral angle (L1-S1), which was approximately 47 degrees. Mean sacral horizontal angle was 35 degrees. From upright to 60 degrees flexion positions, all disc angles decreased uniformly from a total of 53 degrees +/- 6 degrees to 11 degrees +/- 12 degrees in men and 46 degrees +/- 11 degrees to 0 degrees +/- 18 degrees in women. CONCLUSIONS: The dimensions of lumbar vertebral bodies and discs, in upright and flexion positions, appear to be similar between Chinese and Western populations.

Adult↗
PubMed1994

An early molecular response induced by acute overdistension of the rabbit urinary bladder.

In the rabbit, partial urinary bladder outlet obstruction rapidly increases bladder mass and alters several parameters of normal bladder function. Previous experiments on this animal model (Buttyan et al. Neurourol. Urodyn. 11:225-238, 1992) had identified an early molecular response to partial outlet obstruction that involved the profound induction of mRNA encoding a heat-shock gene, hsp-70, as well as induced expression of mRNA for basic fibroblast growth factor and certain protooncogenes. Numerous physiological studies of the hypertrophied rabbit bladder indicate that the primary stimulus for bladder growth may be the initial overdistension that occurs as the bladder fills following partial outlet obstruction. The present study was undertaken to determine if the sequence and characteristics of gene activation during the recovery following a brief period of overdistension of the rabbit bladder are comparable with the gene activity previously described in association with partial outlet obstruction. Rabbit bladders were overdistended to 20% above capacity for 1 hr and then relieved. Bladders were recovered from control (untreated) rabbits and from rabbits at 1 hr or at 1, 3 or 5 days following relief of overdistension. RNAs extracted from these tissues were examined by Northern blot assays for a number of different mRNA transcripts previously shown to be altered by partial outlet obstruction. By 1 hr following the relief from acute overdistension, there was already a greater than 10-fold increase in the expression of hsp-70 related transcripts as well as a marked increase in the expression of mRNA encoding bFGF and decreased expression of TGF-beta 1.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Chloramphenicol acetyltransferase, a cytoplasmic protein is incompatible for export from Bacillus subtilis.

Bacillus subtilis cells expressing a hybrid protein (Lvsss-Cat) consisting of the B. amyloliquefaciens levansucrase signal peptide fused to B. pumilus chloramphenicol acetyltransferase (Cat) are unable to export Cat protein into the growth medium. A series of tripartite protein fusions was constructed by inserting various lengths of the Cat sequences between the levansucrase signal peptide and staphylococcal protein A or Escherichia coli alkaline phosphatase. Biochemical characterization of the various Cat protein fusions revealed that multiple regions in the Cat protein were causing the export defect.

Bacillus↗
PubMed1991

Purification and functional characterization of glutamate-1-semialdehyde aminotransferase from Chlamydomonas reinhardtii.

The formation of delta-aminolevulinic acid, the first committed precursor of chlorophyll biosynthesis, occurs in the chloroplast of plants and algae by the C5-pathway, a three-step, tRNA-dependent transformation of glutamate. Previously, we reported the purification and characterization of the first two enzymes of this pathway, glutamyl-tRNA synthetase and Glu-tRNA reductase from the green alga Chlamydomonas reinhardtii (Chen, M.-W., Jahn, D., Schön, A., O'Neill, G. P., and Söll, D. (1990) J. Biol. Chem. 265, 4054-4057 and Chen, M.-W., Jahn, D., O'Neill, G. P., and Söll, D. (1990) J. Biol. Chem. 265, 4058-4063). Here we present the purification of the third enzyme of the pathway, the glutamate-1-semialdehyde aminotransferase from C. reinhardtii. The enzyme was purified from the membrane fraction of a whole cell extract employing four different chromatographic separations. The apparent molecular mass of the protein was approximately 43,000 Da as analyzed by denaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis, by nondenaturing rate zonal sedimentation on glycerol gradients, and by gel filtration. By these criteria, the enzyme in its active form is a monomer of 43,000 Da. In the presence of pyridoxal 5'-phosphate, purified glutamate-1-semialdehyde aminotransferase converts synthetic glutamate 1-semialdehyde to delta-aminolevulinic acid. The enzyme is inhibited by gabaculine and aminooxyacetate, both typical inhibitors of aminotransferases. The purified glutamate-1-semialdehyde aminotransferase successfully reconstitutes the whole C5-pathway in vitro from glutamate in the presence of purified glutamyl-tRNA synthetase, glutamyl-tRNA reductase, Mg2+, ATP, NADPH, tRNA, and pyridoxal 5'-phosphate.

Aminooxyacetic Acid↗
PubMed1990

Purification and functional characterization of the Glu-tRNA(Gln) amidotransferase from Chlamydomonas reinhardtii.

The formation of glutaminyl-tRNA (Gln-tRNA) in Bacilli, chloroplasts, and mitochondria occurs in a two-step reaction. This involves misacylation of tRNA(Gln) with glutamate by glutamyl-tRNA synthetase and subsequent amidation of Glu-tRNA(Gln) to the correctly acylated Gln-tRNA(Gln) by a specific amidotransferase (Schön, A., Kannangara, C. G., Gough, S., and Söll, D. (1988) Nature 331, 187-190). Here we demonstrate the existence of this pathway in green algae and describe the purification of the Glu-tRNA(Gln) amidotransferase from Chlamydomonas reinhardtii. The purified enzyme showed an Mr of approximately 120,000 when analyzed by glycerol gradient sedimentation and gel filtration. An apparent Mr of 63,000 of the denatured protein was demonstrated by electrophoresis on sodium dodecyl sulfate-polyacrylamide gels. This indicates that the enzyme possesses an alpha 2 structure. The substrate for the purified enzyme is Glu-tRNA(Gln) but not Glu-tRNA(Glu). The enzyme requires ATP, Mg2+, and an amide donor for the conversion. Acceptable amide donors are glutamine, asparagine, and ammonia. Blocking of the glutamine-dependent reaction by alkylation of the protein with 6-diazo-5-oxonorleucine did not inhibit the ammonia-dependent reaction, suggesting that the enzyme has separate glutamine and ammonia binding sites. As suggested by Wilcox (Wilcox, M. (1969) Eur. J. Biochem. 11, 405-412) the amidation reaction may involve glutamyl-phosphate formation, since ATP is cleaved to ADP when the enzyme is incubated with Glu-tRNA(Gln) and ATP. In common with other glutamine amidotransferases, the enzyme also possesses low glutaminase activity. The purified Glu-tRNA(Gln) amidotransferase forms a stable complex with Glu-tRNA(Gln) in the presence of ATP and Mg2+ but in the absence of the amide donor as determined by gradient centrifugation.

Adenosine Triphosphate↗
PubMed1990

Purification and characterization of Chlamydomonas reinhardtii chloroplast glutamyl-tRNA synthetase, a natural misacylating enzyme.

Glutamyl-tRNA synthetase from Chlamydomonas reinhardtii was purified by sequential column chromatography on DEAE-cellulose, phosphocellulose, Mono Q, and Mono S. The apparent molecular mass of the protein when analyzed under both denaturing conditions (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and nondenaturing conditions (rate zonal sedimentation on glycerol gradients) was 62,000 Da; this indicates that the active enzyme is a monomer. The purified glutamyl-tRNA synthetase was identified as the chloroplast enzyme by its tRNA charging specificity. Reversed-phase chromatography of unfractionated C. reinhardtii tRNA resolved four peaks of glutamate acceptor RNA when assayed with the purified enzyme. The enzyme can also glutamylate Escherichia coli tRNA(2Glu), but not cytoplasmic tRNA(Glu) from yeast or barley. In addition, the enzyme misacylates chloroplast tRNA(Gln) with glutamate. A similar mischarging phenomenon has been demonstrated for the barley chloroplast enzyme (Schön, A., Kannangara, C.G., Gough, S., and Söll, D. (1988) Nature 331, 187-190) and for Bacillus subtilis glutamyl-tRNA synthetase (Proulx, M., Duplain, L., Lacoste, L., Yaguchi, M., and Lapointe, J. (1983) J. Biol. Chem. 258, 753-759).

Acylation↗
PubMed1990

Purification of the glutamyl-tRNA reductase from Chlamydomonas reinhardtii involved in delta-aminolevulinic acid formation during chlorophyll biosynthesis.

The formation of delta-aminolevulinic acid, the first committed precursor in porphyrin biosynthesis, occurs in certain bacteria and in the chloroplasts of plants and algae in a three-step, tRNA-dependent transformation of glutamate. Glutamyl-tRNA reductase, the second enzyme of this pathway, reduces the activated carboxyl group of glutamyl-tRNA (Glu-tRNA) in the presence of NADPH and releases glutamate 1-semialdehyde (GSA). We have purified Glu-tRNA reductase from Chlamydomonas reinhardtii by employing six different chromatographic separations. The apparent molecular mass of the protein when analyzed under both denaturing (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and nondenaturing conditions (rate zonal sedimentation on glycerol gradients) was 130,000 Da; this indicates that the active enzyme is a monomer. In the presence of NADPH Glu-tRNA reductase catalyzed the reduction to GSA of glutamate acylated to the homologous tRNA. Thus, the reductase alone is sufficient for conversion of Glu-tRNA to GSA. In the absence of NADPH, a stable complex of Glu-tRNA reductase with Glu-tRNA can be isolated.

Aminolevulinic Acid↗
PubMed1989

delta-Aminolevulinic acid biosynthesis in Escherichia coli and Bacillus subtilis involves formation of glutamyl-tRNA.

Cell-free extracts of Escherichia coli and Bacillus subtilis catalyzed the tRNA-dependent, RNase A-sensitive formation of delta-aminolevulinic acid (ALA) from glutamate. Cell extracts prepared from cultures of E. coli grown under aerobic or anaerobic conditions had similar levels of ALA biosynthetic activity. Both the tRNA-stimulated conversion of glutamate to ALA and the conversion of glutamate-1-semialdehyde to ALA were inhibited by gabaculin. However, gabaculin had no effect on the growth of either E. coli or B. subtilis. The tRNA-dependent transformation of glutamate to ALA in E. coli and B. subtilis thus appears to be very similar to the pathway found in cyanobacteria, certain obligate anaerobic eubacteria, archaebacteria and in the chloroplasts of algae and higher plant species.

Alanine↗
PubMed1988

Formation of the chlorophyll precursor delta-aminolevulinic acid in cyanobacteria requires aminoacylation of a tRNAGlu species.

In the chloroplasts of higher plants and algae, the biosynthesis of the chlorophyll precursor delta-aminolevulinic acid (ALA) involves at least three enzymes and a tRNA species. Here we demonstrate that in cell extracts of the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 ALA was formed from glutamate in a series of reactions in which activation of glutamate by glutamyl-tRNAGlu formation was the first step. The activated glutamate was reduced by a dehydrogenase which displayed tRNA sequence specificity. Fractionation of strain 6803 tRNA by reverse-phase chromatography and polyacrylamide gel electrophoresis yielded two pure tRNAGlu species which stimulated ALA synthesis in vitro. These tRNAs had identical primary sequences but differed in the nucleotide modification of their anticodon. The 6803 tRNAGlu was similar to the sequences of tRNAGlu species or tRNAGlu genes from Escherichia coli and from chloroplasts of Euglena gracilis and higher plants. Southern blot analysis revealed at least two tRNAGlu gene copies in the 6803 chromosome. A glutamate-1-semialdehyde aminotransferase, the terminal enzyme in the conversion of glutamate to ALA in chloroplasts, was detected in 6803 cell extracts by the conversion of glutamate-1-semialdehyde to ALA and by the inhibition of this reaction by gabaculin.

Aminolevulinic Acid↗
PubMed1984

Equilibria in 5-S ribosomal RNA secondary structure. Bulges and interior loops in 5-S RNA secondary structure may serve as articulations for a flexible molecule.

The basic assumption in this paper is that the secondary structure of a 5-S ribosomal RNA cannot be represented by a single model. We propose that the molecule can adopt, at least within the ribosome, a series of slightly different structures of nearly equal stability. The different structures arise from the existence of ambiguous base-pairing opportunities in bulged helices and the adjacent interior loops. In eubacterial 5-S RNAs there is one such an area, in eukaryotic 5-S RNAs two such areas that can give rise to structural switches. We explain how a change in secondary structure in these areas may influence the relative orientation of the surrounding helices, in other words how bulges and interior loops may serve as articulations and give rise to a flexible tertiary structure.

Animals↗
PubMed1984

The nucleotide sequences of the 5 S rRNAs of seven molds and a yeast and their use in studying ascomycete phylogeny.

The sequences of the 5 S rRNAs isolated from 8 ascomycete species belonging to the genera Aspergillus, Penicillium, Acremonium and Candida are reported. Two of the examined strains each yielded a mixture of 3 slightly different 5 S RNAs, which were individually sequenced after fractionation. A previously published sequence for Aspergillus nidulans 5 S RNA was found to contain errors. Reconstruction of an evolutionary tree based on 5 S RNA sequences showed that the 16 presently examined ascomycetes form three clusters. The same threefold partition can be observed in the secondary structure pattern, each cluster showing a slightly different variant of the general 5-helix model for 5 S rRNA (De Wachter, Chen and Vandenberghe (1982) Biochimie 64, 311-329), and different sets of secondary structure equilibrium forms in helices C and E of the aforementioned model.

Aspergillus flavus↗
PubMed1982

Conservation of secondary structure in 5 S ribosomal RNA: a uniform model for eukaryotic, eubacterial, archaebacterial and organelle sequences is energetically favourable.

The most commonly accepted secondary structure models for 5S RNA differ for molecules of eubacterial origin, where the four-helix model of Fox and Woese is generally cited, and those of eukaryotic origin, where a fifth helix is assumed to exist. We have carefully aligned all available sequences from eukaryotes, eubacteria, chloroplasts, archaebacteria and plant mitochondria. We could thus derive a unified secondary structure model applicable to all 5S RNA sequences known to-date. It contains the five helices already present in the eukaryotic model, extended by additional segments that were not previously assumed to be universally present. One of the helices can be written in two equilibrium forms, which could reflect the existence of a flexible, dynamic structure. For the derivation of the model and the estimation of the free energies we followed a set of rules optimized to predict the tRNA cloverleaf. The stability of the unified model is higher than that of nearly all previously proposed sequence-specific and general models.

Animals↗