Localization of N-Terminal Sequences in Human AMP Deaminase Isoforms That Influence Contractile Protein Binding

Mahnke-Zizelman, Donna K.; Sabina, Richard L.

doi:10.1006/bbrc.2001.5180

Cited by 20 publications

(15 citation statements)

References 33 publications

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“…Recombinant virus was used to infect 12 confluent T-185 flasks of Sf9 cells and recombinant enzymes were purified by phosphocellulose chromatography using a previously described protocol (18). Unless otherwise stated, leupeptin was included in all extraction and storage buffers to minimize N-terminal proteolysis, as previously described (19).…”

Section: Methodsmentioning

confidence: 99%

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Han

Bingman

Mahnke

et al. 2006

Journal of Biological Chemistry

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Embryonic factor 1 (FAC1) is one of the earliest expressed plant genes and encodes an AMP deaminase (AMPD), which is also an identified herbicide target. This report identifies an N-terminal transmembrane domain in Arabidopsis FAC1, explores subcellular fractionation, and presents a 3.3-Å globular catalytic domain x-ray crystal structure with a bound herbicide-based transition state inhibitor that provides the first glimpse of a complete AMPD active site. FAC1 contains an (␣/␤) 8 -barrel characterized by loops in place of strands 5 and 6 that places it in a small subset of the amidohydrolase superfamily with imperfect folds. Unlike tetrameric animal orthologs, FAC1 is a dimer and each subunit contains an exposed Walker A motif that may be involved in the dramatic combined K m (25-80-fold lower) and V max (5-6-fold higher) activation by ATP. Normal mode analysis predicts a hinge motion that flattens basic surfaces on each monomer that flank the dimer interface, which suggests a reversible association between the FAC1 globular catalytic domain and intracellular membranes, with N-terminal transmembrane and disordered linker regions serving as the anchor and attachment to the globular catalytic domain, respectively.Embryonic factor 1 (FAC1) 5 was recently identified as one of the earliest expressed plant genes and is essential for the zygote to embryo transition in Arabidopsis thaliana (1). The zygote-lethal phenotype is characterized by developmental arrest at the 8 -16-cell stage and mutant embryo shriveling 2-3 days after fertilization. The Arabidopsis FAC1 locus encodes an AMP deaminase (AMPD; EC 3.5.4.6), which is a eukaryotic enzyme that catalyzes the hydrolytic deamination of AMP to IMP. AMPD has also been identified as the intracellular target for a class of herbicides that are produced by fungal pathogens. Carbocyclic coformycin was initially discovered in Saccharothrix (2), and plant cells can take up this diffusible nucleoside and 5Ј-phosphorylate it to produce a potent transition state inhibitor of AMPD (3). Exposure to carbocyclic coformycin results in cessation of seedling growth, followed by paling and necrosis at the apical meristem (3). Coformycin, a structurally related compound produced by a number of microbes (4, 5), also has herbicidal properties (5). Although the intracellular metabolism of this compound in plants has not been examined, its mode of action is presumably similar because coformycin 5Ј-phosphate is a transition state inhibitor of rabbit muscle AMPD (6). Both herbicides are inhibitors of mammalian adenosine deaminase (ADA) (3, 6), but the lack of this enzyme in plants (3, 7-9) supports the argument that AMPD is the intracellular target for their nucleotide derivatives. Taken together, these observations suggest that AMPD is essential throughout the plant life cycle. However, the underlying basis for lethality of a FAC1 null phenotype and for herbicidal toxicity related to the catalytic inhibition of this enzyme is not known.AMPD catalyzes the initial step in adenine to guanine ribon...

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Section: Methodsmentioning

confidence: 99%

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Han

Bingman

Mahnke

et al. 2006

Journal of Biological Chemistry

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“…Mammalian AMPD undergoes limited proteolysis in vitro; the degradation is limited to the N-terminal regions of the AMPD isozymes. Proteolysis of the N-terminal fragments does not reduce significantly catalytic activity of the enzyme (Mahnke-Zizelman and Sabina 2001 ). The use of new recombinant technologies allowed to synthesize full-size AMPD proteins with intact N-terminal fragments (Sabina et al 1984 ).…”

Section: Discussionmentioning

confidence: 99%

“…This is normally observed during purification of the enzyme and its further storage. The molecular mass of AMPD1 subunit isolated freshly from autopsied human skeletal muscle (60–72 kDa) (Mahnke-Zizelman and Sabina 2001 ; Stankiewicz 1981 ) differs markedly from that predicted on the basis of cDNA sequencing (86–87 kDa) (Sabina et al 1984 ). This discrepancy is most probably a result of proteolysis taking place during the process of purification (Sabina et al 1984 ).…”

Section: Discussionmentioning

confidence: 99%

Isozymes of AMP-Deaminase in Muscles Myasthenia Gravis Patients

Rybakowska

Bakuła

Kaletha

2016

Int J Pept Res Ther

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Similar symptoms observed in Myasthenia gravis (MG) can be also detected in the case of skeletal muscle AMP-deaminase deficiency. We compared the activity and expression of AMP-deaminase (AMPD) products in skeletal muscles of MG patients and MG-free individuals. The activity of AMP-deaminase in the muscles of MG patients was significantly higher than in the controls and was 2.05 µmol/min/mg protein (±0.31). The two groups differ in level of AMPD product expression. Furthermore in MG-group molecular size of isoform AMPD1 is 90 kDa in contrast to MG-free group where is present 70 kDa isoform of enzyme. The data suggests that the disturbances in transmission of neuronal signaling, taking place in the skeletal muscles of MG patients, may also change energetic metabolism of the affected muscles by changing molecular mass of isoform.

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“…A rabbit polyclonal antibody against MX1 was generously provided by Dr Otto Haller (Institut fu¨r Medizinische Mikrobiologie und Hygiene, Universita¨t Freiburg, Freiburg, Germany) and was utilized at a 1:250 dilution (Engelhardt et al, 2004). A rabbit polyclonal anti-AMPD3 antiserum was generously provided by Dr Richard L Sabina (Medical College of Wisconsin, Milwaukee, WI, USA) and was used at a 1:150 dilution (Mahnke-Zizelman and Sabina, 2000). Because of difficulty in detecting DN-Akt protein, a combination of three primary antibodies was used for Western blotting of total Akt: anti-Akt (catalog # P8103S, New England Biolabs (Beverly, MA, USA), 1:200) plus anti-Akt (#MAB17751, R&D Systems Inc. (Minneapolis, MN, USA), 1:200) plus anti-Akt (clone 302407, R&D Systems, 1:200).…”

Section: Western Blottingmentioning

confidence: 99%

Effect of Akt inhibition on scatter factor-regulated gene expression in DU-145 human prostate cancer cells

Gao

Fan

et al. 2006

Oncogene

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The cytokine scatter factor (SF) (hepatocyte growth factor) transduces various biologic actions, including cell motility, invasion, angiogenesis and apoptosis inhibition. The latter is relevant to understanding the role of SF in promoting tumor cell survival in different contexts, for example, detachment from basement membrane, growth in metastatic sites and responses to chemo-and radiotherapy. Previously, we showed that SF protects cells against apoptosis owing to DNA damage, by a mechanism involving phosphoinositol-3-kinase/c-Akt signaling. Here, we used DNA microarray assays to identify c-Aktregulated genes that might contribute to cell protection. DU-145 human prostate cancer cells were transfected7a dominant-negative mutant Akt, treated7SF and analysed for gene expression using Affymetrix arrays. These studies identified SF-regulated genes for which induction was c-Akt-dependent vs -independent. Selected microarray findings were confirmed by semiquantitative and quantitative reverse transcription-polymerase chain reaction. We tested the contribution of four SF-inducible/ c-Akt-dependent genes (AMPD3, EPHB2, MX1 and WNT4) to protection against adriamycin (a DNA topoisomerase IIa inhibitor) using RNA interference. Knockdown of each gene except EPHB2 caused a small but significant reduction in the SF cell protection. The lack of effect of EPHB2 knockdown may be due to the fact that DU-145 cells contain a single-mutant EPHB2 allele. A combination of three small interfering RNAs blocked most of the protection by SF in both DU-145 and T47D cells. These findings identify novel c-Akt-regulated genes, some of which contribute to SF-mediated cytoprotection.

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Localization of N-Terminal Sequences in Human AMP Deaminase Isoforms That Influence Contractile Protein Binding

Cited by 20 publications

References 33 publications

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Isozymes of AMP-Deaminase in Muscles Myasthenia Gravis Patients

Effect of Akt inhibition on scatter factor-regulated gene expression in DU-145 human prostate cancer cells

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