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Biomedical subjects

M S Anderson

Publications and source records attributed to M S Anderson.

At least 55 records · Page 3Linked to original sources
PubMed1993

The firA gene of Escherichia coli encodes UDP-3-O-(R-3-hydroxymyristoyl)-glucosamine N-acyltransferase. The third step of endotoxin biosynthesis.

The possibility that the firA gene of Escherichia coli (Dicker, I. B., and Seetharam, S. (1991) Mol. Microbiol. 6, 817-823) might function in lipid A biosynthesis was examined based on its homology to the lpxA gene, which encodes UDP-N-acetylglucosamine O-acyl-transferase, the first enzyme in lipid A formation. Extracts of a temperature-sensitive firA mutant, RL-25, were assayed for their ability to acylate UDP-GlcNAc, using a coupled assay. The results suggested that extracts of RL-25 might be defective in the third enzyme of this pathway, the UDP-3-O-(R-3-hydroxymyristoyl)-glucosamine N-acyltransferase. Living cells of RL-25 also displayed a 5-fold decreased rate of lipid A biosynthesis at the nonpermissive temperature as judged by a 32Pi incorporation assay. In order to examine N-acyltransferase activity directly, the substrate [alpha-32P]UDP-3-O-(R-3-hydroxymyristoyl)-GlcN was synthesized enzymatically. N-Acyltransferase specific activity in RL-25 extracts was reduced to less than 10% of wild-type. When the wild-type firA gene was cloned into a T7-based expression vector, N-acyltransferase specific activity increased almost 360-fold relative to wild-type extracts, demonstrating that firA is the structural gene for the enzyme. The N-acyltransferase displays absolute specificity for the R-3-OH moiety of R-3-hydroxymyristoyl-ACP, as does the O-acyltransferase, consistent with the placement of R-3-hydroxymyristate in E. coli lipid A.

Acyltransferases↗
PubMed1993

The chondroitin sulfate form of invariant chain can enhance stimulation of T cell responses through interaction with CD44.

Invariant chain (Ii) is a nonpolymorphic glycoprotein that associates with major histocompatibility complex class II molecules and has been shown to mediate several functions in class II-restricted antigen presentation. A small proportion of Ii is modified by the addition of chondroitin sulfate (Ii-CS), and this form of Ii is associated with class II on the surface of antigen-presenting cells. In this report we show that expression of Ii-CS dramatically enhanced the ability of class II-positive EL4 transfectants to stimulate class II-dependent allogeneic and mitogenic T cell responses. Antibody blocking studies and the ability of CD44 to bind directly to Ii-CS suggest that Ii-CS can function as an accessory molecule during T cell responses through interactions with CD44.

Animals↗
PubMed1993

Enhancement of merocyanine 540-mediated phototherapy by salicylate.

Merocyanine 540 (MC540) is a photosensitizing dye of potential use in the purging of cancer cells from autologous bone marrow explants. Treatment of marrow with MC540, followed by illumination with visible light, selectively kills neoplastic cells while sparing a sufficient number of stem cells to allow marrow engraftment. The photodynamic action of MC540 is thought to be mediated by reactive oxygen species, particularly singlet oxygen. We have previously shown that salicylic acid (SA) scavenges MC540-generated singlet oxygen. In this work, we sought to abrogate MC540-mediated cell killing of murine L1210 and human K562 leukemia cells with salicylate. Paradoxically, the presence of salicylate during illumination in the presence of MC540 appreciably enhanced cell killing. Enhancement was dependent on salicylate concentration in the range 0.1 to 10 mM, with 1.0 mM SA potentiating the MC540-mediated reduction in survival of L1210 and K562 cells by factors of 2.7 and 1.9, respectively. Neither preincubation with SA followed by washing prior to illumination nor addition of SA following illumination altered MC540-mediated cell killing, indicating that potentiation was dependent on the presence of SA during illumination. Illumination in the presence of salicylate alone did not diminish cell viability. In addition to SA, a number of structurally related compounds including dihydroxybenzoic acids, aspirin, and sodium benzoate also enhanced MC540-mediated cell killing. Potentiation of leukemic cell killing by salicylate could provide a basis for enhancing the clinical efficacy of MC540-mediated phototherapy.

Animals↗
PubMed1992

Invariant chain can function as a chaperone protein for class II major histocompatibility complex molecules.

During biosynthesis, class II major histocompatibility complex molecules are intimately associated with invariant chain (Ii). The Ii-class II association has been shown to block peptide-class II binding and to affect the ultimate conformation of class II expressed on the cell surface. To assess the biochemical basis for the effects of Ii on class II, we have analyzed the biosynthesis of class II in EL4 cells transfected with I-Ad with and without Ii. In these studies, we found that Ii had a profound effect on the biosynthesis of I-Ad. In the absence of Ii, class II could form dimers efficiently, but these dimers appeared to be misfolded and this altered conformation resulted in the loss of some monoclonal antibody epitopes and inefficient transport from the endoplasmic reticulum to the Golgi. In addition, class II that was transported through the Golgi accumulated an abnormally increased molecular mass associated with N-linked glycosylation. Subsequent transfection of Ii into these cells resulted in recovery of normal class II conformation, causing a restoration of monoclonal antibody epitopes, efficient intracellular transport, and normal glycosylation. Together, these data indicate that Ii can have a profound effect on the folding, transport, and modification of class II molecules and suggest that one function of Ii may be to act as a class II-specific chaperone.

Animals↗
PubMed1992

Possible influences on the expression of X chromosome-linked dystrophin abnormalities by heterozygosity for autosomal recessive Fukuyama congenital muscular dystrophy.

Abnormalities of dystrophin, a cytoskeletal protein of muscle and nerve, are generally considered specific for Duchenne and Becker muscular dystrophy. However, several patients have recently been identified with dystrophin deficiency who, before dystrophin testing, were considered to have Fukuyama congenital muscular dystrophy (FCMD) on the basis of clinical findings. Epidemiologic data suggest that only 1/3500 males with autosomal recessive FCMD should have abnormal dystrophin. To explain the observation of 3/23 FCMD males with abnormal dystrophin, we propose that dystrophin and the FCMD gene product interact and that the earlier onset and greater severity of these patients' phenotype (relative to Duchenne muscular dystrophy) are due to their being heterozygous for the FCMD mutation in addition to being hemizygous for Duchenne muscular dystrophy, a genotype that is predicted to occur in 1/175,000 Japanese males. This model may help explain the genetic basis for some of the clinical and pathological variability seen among patients with FCMD, and it has potential implications for understanding the inheritance of other autosomal recessive disorders in general. For example, sex ratios for rare autosomal recessive disorders caused by mutations in proteins that interact with X chromosome-linked gene products may display predictable deviation from 1:1.

Abnormalities, Multiple↗
PubMed1992

The molecular and biochemical basis of Duchenne muscular dystrophy.

Duchenne and Becker muscular dystrophy (DMD, BMD) have both been clinically recognized for over 100 years, yet throughout much of that time nothing beyond clinical evaluation and supportive care during the disease course was available to patients. The identification of the molecular basis of DMD/BMD in 1986 paved the way for extensive progress toward the understanding, diagnosis and treatment of this disease.

Chromosome Mapping↗
PubMed1992

The epidemiology of myasthenia gravis in central and western Virginia.

We conducted a study of the epidemiology of myasthenia gravis (MG) in four locations in central and western Virginia from 1970 through 1984. The population surveyed was 555,851 in 1984. A total of 73 new cases of MG occurred during the survey period, producing an overall average annual incidence rate of 9.1 per million. The point prevalence rate in 1980 was 13.4 per 100,000, and in 1984 it was 14.2. Approximately 15% of the population was black, and we found that incidence and prevalence rates for the black population were higher than the corresponding white population. When the population was subdivided into <50 and 50+ age groups, the incidence and prevalence were significantly higher in the older group. The rates we report here are higher than rates reported from any other locality. The reasons for the higher rates include optimal case identification, survey of a population with a higher incidence, and increasing aging of the population.

Adult↗
PubMed1991

Acyl-acyl carrier protein specificity of UDP-GlcNAc acyltransferases from gram-negative bacteria: relationship to lipid A structure.

Lipid A, the component of lipopolysaccharide that provides the membrane anchor of the core and O-antigen sugars, is known to contain characteristic R-3-hydroxy fatty acids bound to the 2,2' (N-linked) and 3,3' (O-linked) positions of the glucosamine disaccharide in different gram-negative bacteria. The studies reported here show that it is the acyl-acyl carrier protein specificities of the enzymes UDP-GlcNAc-O-acyltransferase and UDP-3-O-[(R)-3-hydroxyacyl]-GlcN-N-acyltransferase that determine the nature of these fatty acids.

Acinetobacter↗
PubMed1991

Plasma membrane properties regulating the sensitivity of leukemia, lymphoma, and solid tumor cells to merocyanine 540-sensitized photoirradiation.

Merocyanine 540 (MC 540) is a photosensitizing dye that has been used in a phase I clinical trial for the purging of leukemia and lymphoma cells from autologous bone marrow grafts. In this paper we examine the role of plasma membrane negative charge, plasma membrane fluidity, and plasma membrane hydrophobicity in the regulation of a cell's susceptibility to MC 540-sensitized photoirradiation. Among solid tumor cells, we found an inverse correlation between surface electronegativity, affinity for dye molecules, and susceptibility to MC 540-sensitized photoinactivation. That is, the least electronegative cells bound the highest amount of dye and were the most susceptible to dye-sensitized photoirradiation. By contrast, no such correlations were found among leukemia/lymphoma cells. This suggested that dye binding and susceptibility to MC 540-mediated photodynamic damages are regulated differently in hematopoietic/lymphopoietic and solid tumor cells.

Bone Marrow↗
PubMed1989

Isopentenyl diphosphate:dimethylallyl diphosphate isomerase. An improved purification of the enzyme and isolation of the gene from Saccharomyces cerevisiae.

Isopentenyl diphosphate:dimethylallyl diphosphate isomerase (IPP isomerase) is an enzyme in the isoprenoid biosynthetic pathway which catalyzes the interconversion of the primary five-carbon homoallylic and allylic diphosphate building blocks. We report a substantially improved procedure for purification of this enzyme from Saccharomyces cerevisiae. An amino-terminal sequence (35 amino acids) was obtained from a highly purified preparation of IPP isomerase. Oligonucleotide probes based on the protein sequence were used to isolate the structural gene encoding IPP isomerase from a yeast lambda library. The cloned gene encodes a 33,350-dalton polypeptide of 288 amino acids. A 3.5-kilobase EcoRI fragment containing the gene was subcloned into the yeast shuttle vector YRp17. Upon transformation with plasmids containing the insert, a 5-6-fold increase in IPP isomerase activity was seen in transformed cells relative to YRp17 controls, confirming the identity of the cloned gene. This is the first reported isolation of the gene for IPP isomerase.

Amino Acid Sequence↗
PubMed1989

Farnesyl diphosphate synthetase. Molecular cloning, sequence, and expression of an essential gene from Saccharomyces cerevisiae.

Farnesyl diphosphate (FPP) synthetase is a key enzyme in isoprenoid biosynthesis which supplies C15 precursors for several classes of essential metabolites including sterols, dolichols, and ubiquinones. The structural gene for FPP synthetase was isolated on a 4.5-kilobase EcoRI genomic restriction fragment from the yeast Saccharomyces cerevisiae. The clone encodes a 40,483-dalton polypeptide of 342 amino acids with a high degree of similarity to the protein encoded by a putative rat liver clone of FPP synthetase (Clarke, C. F., Tanaka, R. D., Svenson, K., Wamsley, M., Fogelman, A. M., and Edwards, P. A. (1987) Mol. Cell Biol. 7, 3138-3146) and to an active site protein fragment from avian liver FPP synthetase (Brems, D. N., Bruenger, E., and Rilling, H. C. (1981) Biochemistry 20, 3711-3718). When cloned into the yeast shuttle vector YRp17, the 4.5-kilobase EcoRI fragment directed a 2-3-fold over-expression of FPP synthetase activity in transformed yeast cells. The levels of expression were independent of culture growth phase and orientation of the insert, indicative of a functional promoter in the clone. Disruption of the FPP synthetase gene from a diploid yeast strain, followed by dissection and analysis of tetrads, demonstrates that the gene is an essential, single copy number gene in yeast. The gene for FPP synthetase resides on chromosome XI as judged from Southern blots of separated yeast chromosomes.

Amino Acid Sequence↗
PubMed1989

Priorities for nutrition content in a medical school curriculum: a national consensus of medical educators.

The ASCN Committee on Medical/Dental School and Residency Nutrition Education conducted a series of activities to establish guidelines for nutrition core content in a medical school curriculum. These activities included mail surveys of medical-nutrition educators and a representative group of medical school curriculum administrators and a national consensus workshop of nutrition educators. Results indicated close agreement between the nutrition educators and curriculum administrators (r = 0.89, p less than 0.0001) on the importance ratings of 41 nutrition topics and on the number of hours of nutrition course work that medical schools should provide (44 vs 37 h, respectively, p = 0.14). There was consensus among the nutrition educators that 26 topics should be given priority ratings as essential for inclusion in medical course work. Further prioritization of these topics resulted in a listing of core content topics and subtopics to serve as a guide to administrators and educators for planning nutrition course work in a medical school curriculum.

Curriculum↗
PubMed1988

Biosynthesis of lipid A in Escherichia coli: identification of UDP-3-O-[(R)-3-hydroxymyristoyl]-alpha-D-glucosamine as a precursor of UDP-N2,O3-bis[(R)-3-hydroxymyristoyl]-alpha-D-glucosamine.

The lipid A disaccharide of the Escherichia coli envelope is synthesized from the two fatty acylated glucosamine derivatives UDP-N2,O3-bis[(R)-3-hydroxytetradecanoyl]-alpha-D- glucosamine (UDP-2,3-diacyl-GlcN) and N2,O3-bis[(R)-3-hydroxytetradecanoyl]-alpha-D-glucosamine 1-phosphate (2,3-diacyl-GlcN-1-P) [Ray, B. L., Painter, G., & Raetz, C. R. H. (1984) J. Biol. Chem. 259, 4852-4859]. We have previously shown that UDP-2,3-diacyl-GlcN is generated in extracts of E. coli by fatty acylation of UDP-GlcNAc, giving UDP-3-O-[(R)-3-hydroxymyristoyl]-GlcNAc as the first intermediate, which is rapidly converted to UDP-2,3-diacyl-GlcN [Anderson, M. S., Bulawa, C. E., & Raetz, C. R. H. (1985) J. Biol. Chem. 260, 15536-15541; Anderson, M. S., & Raetz, C. R. H. (1987) J. Biol. Chem. 262, 5159-5169]. We now demonstrate a novel enzyme in the cytoplasmic fraction of E. coli, capable of deacetylating UDP-3-O-[(R)-3-hydroxymyristoyl]-GlcNAc to form UDP-3-O-[(R)-3-hydroxymyristoyl]glucosamine. The covalent structure of the previously undescribed UDP-3-O-[(R)-3-hydroxymyristoyl] glucosamine intermediate was established by 1H NMR spectroscopy and fast atom bombardment mass spectrometry. This material can be made to accumulate in E. coli extracts upon incubation of UDP-3-O-[(R)-3- hydroxymyristoyl]-GlcNAc in the absence of the fatty acyl donor [(R)-3-hydroxymyristoyl]-acyl carrier protein. However, addition of the isolated deacetylation product [UDP-3-O-[(R)-3-hydroxymyristoyl] glucosamine] back to membrane-free extracts of E. coli in the presence of [(R)-3-hydroxymyristoyl]-acyl carrier protein results in rapid conversion of this compound into the more hydrophobic products UDP-2,3-diacyl-GlcN, 2,3-diacyl-GlcN-1-P, and O-[2-amino-2-deoxy-N2,O3- bis[(R)-3-hydroxytetradecanoyl]-beta-D-glucopyranosyl]-(1----6)-2-amino- 2-deoxy-N2,O3-bis[(R)-3-hydroxytetradecanoyl]-alpha-D- glucopyranose 1-phosphate (tetra-acyldisaccharide-1-P), demonstrating its competency as a precursor. In vitro incubations using [acetyl-3H]UDP-3-O-[(R)-3-hydroxymyristoyl]-GlcNAc confirmed release of the acetyl moiety in this system as acetate, not as some other acetyl derivative. The deacetylation reaction was inhibited by 1 mM N-ethylmaleimide, while the subsequent N-acylation reaction was not. Our observations provide strong evidence that UDP-3-O-[(R)-3-hydroxymyristoyl]glucosamine is a true intermediate in the biosynthesis of UDP-2,3-diacyl-GlcN and lipid A.

Acetic Anhydrides↗
PubMed1988

Identification of the neuropeptide transmitter proctolin in Drosophila larvae: characterization of muscle fiber-specific neuromuscular endings.

The cellular localization of the peptide neurotransmitter proctolin was determined for larvae of the fruitfly Drosophila melanogaster. Proctolin was recovered from the CNS, hindgut, and segmental bodywall using reverse-phase HPLC, and characterized by bioassay, immunoassay, and enzymatic analysis. A small, stereotyped population of proctolin-immunoreactive neurons was found in the larval CNS. Several of the identified neurons may be efferents. In the periphery, proctolin-immunoreactive neuromuscular endings were identified on both visceral and skeletal muscle fibers. On the hindgut, the neuropeptide is associated with endings on intrinsic circular muscle fibers. We propose that the hindgut muscle fibers are innervated by central neurons homologous to previously described proctolinergic efferents of grasshoppers. The segmental bodywall innervation consists of a pattern of segment-specific junctions on several singly identifiable muscle fibers. While it is generally accepted that Drosophila muscle fibers are innervated by glutamatergic motoneurons, our data indicate that a specialized subset of muscle fibers are also innervated by peptidergic efferents.

Abdomen↗